bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5113: CXCL12/CXCR4 and integrin signal activation by cancer-associated fibroblasts enhances the chemoresistance in gastric cancer
Authors: Tamaoki Yuka , Ishimoto Takatsugu , Miyake Keisuke , Izumi Daisuke , Kuroda Daisuke , Eto Tsugio , Arima Kota , Shigaki Hironobu , Kurashige Junji , Iwatsuki Masaaki , Baba Yoshifumi , Sakamoto Yasuo , Yoshida Naoya , Baba Hideo
Published: July 2016

Purpose: Gastric cancer (GC) is an aggressive cancer metastasizing with a high propensity for peritoneal dissemination and liver metastasis. Some studies have been shown that not only cancer cells but also their surrounding stroma, especially cancer-associated fibroblasts (CAFs) play an important role in tumor development. Previously, we reported that CXCL12-CXCR4 and integrin β1 signal activation by CAFs co-operates for promoting FAK phosphorylation and enhances the invasiveness of GC cells in extracellular matrix (ECM).On the other hand, a previous study presented that CXCR4 and integrin signaling co-operates in mediating adhesion and chemoresistance in small cell lung cancer cells. Therefore, we hypothesized that CXCL12-CXCR4 and integrin signal activation by CAFs enhances the chemoresistance in GC as well. The aim of current study is to elucidate the significance of CAFs mediating signal activation for chemoresistance in GCs.

Experimental Design: We first performed chemo-sensitive assays using GC cells cultured with fresh medium or with CAF-conditioned-medium (CM), and these experiments were conducted on non-coated and Matrigel-coated plates. Next, we examined the CXCL12-CXCR4 and integrin related signaling by western blotting analysis. Furthermore, we investigated the critical molecule for chemoresistance using silencing CXCR4 or integrin β1 in GC cells.

Results: Chemo-sensitive assays revealed that GC cells cultured with CAF-CM showed more resistant to cisplatin and reactive oxygen spices (ROS) than those cultured with fresh medium. Moreover, GC cells with CAF-CM on Matrigel-coated plates exhibited remarkable resistance in these assays. On the other hand, western blotting analysis showed that Akt activity in GC cells cultured with CAF-CM was up-regulated in concurrence with FAK phosphorylation. Finally, the chemoresitance induced by CAFs was significantly suppressed by CXCR4 or integrin β1 silencing.

Conclusions: These results suggest that CXCL12/CXCR4 and integrinβ1 signal activation by CAFs promoted the chemoresistance through enhancing their interaction with ECM in GC. This mechanism underlying the chemoresistance may provide a novel therapeutic target in advanced GCs.

Citation Format: Yuka Tamaoki, Takatsugu Ishimoto, Keisuke Miyake, Daisuke Izumi, Daisuke Kuroda, Tsugio Eto, Kota Arima, Hironobu Shigaki, Junji Kurashige, Masaaki Iwatsuki, Yoshifumi Baba, Yasuo Sakamoto, Naoya Yoshida, Hideo Baba. CXCL12/CXCR4 and integrin signal activation by cancer-associated fibroblasts enhances the chemoresistance in gastric cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5113.
Abstract 5134: Evaluation of immune tolerance mechanisms in pulmonary squamous cell and adenocarcinomas - how to interfere beyond PDL1 blockade
Authors: Popper Helmut H. , Brcic Luka , Absenger Gudrun , Prietl Barbara , Lindenmann Joerg , Balic Marija
Published: July 2016

Introduction: Immunotherapy has emerged as a new tool to treat patients with NSCLC. Programmed death ligand 1 (PDL1) expressed on carcinoma cells serve as a marker predicting the response of anti-PDL antibody therapy. However, the results of clinical studies are conflicting as patients might profit from therapy despite not expressing PDL1 on their tumors, and also some tumors expressing PDL1 do not respond to therapy. It is evident that NSCLC cells use other mechanisms of immunotolerance.

Methods: Surgically removed specimen of lung cancer are evaluated for infiltrating innate as well as specific immune cell in tumor centers and invasive front. In addition bronchoalveolar lavage is taken simultaneously and evaluated by flow cytometry for the same immune cells as the tissues. These cell types are evaluated: dendritic cells (classical, plasmocytoid, monocytoid), macrophages (M1 and M2), lymphocytes of Bcell lineage, lymphocytes of Tcell lineage, regulatory Tcells, natural killer cells. In addition tumor cells and lymphocytes are studied for their expression of PD1, PDL1, and PDL2, respectively

Results and discussion: Within lung cancer several different mechanisms of inducing immune tolerance emerged: PD1-PDL1 interaction of tumor cells and CD8 lymphocytes, plasmocytoid as well as monocytoid dendritic cell induced downregulation of cytotoxic Tcell action, influx of myeloid derived cells inducing immune tolerance against tumor cells, M2 macrophage polarization with enhancement of stroma remodeling for tumor cell invasion and reduced Tcell reaction, increase of Treg cells inducing immune tolerance. Some of these mechanisms are seen in combination with PD1-PDL1 induced immune tolerance. This might explain, why some patients do not benefit from anti-PDL1 therapy. BAL emerged as a powerful tool in the evaluation of the immune reaction in NSCLC patients and might be a tool for evaluation of immune responses as well as a diagnostic tool to evaluate resistance mechanisms.

Citation Format: Helmut H. Popper, Luka Brcic, Gudrun Absenger, Barbara Prietl, Joerg Lindenmann, Marija Balic. Evaluation of immune tolerance mechanisms in pulmonary squamous cell and adenocarcinomas - how to interfere beyond PDL1 blockade. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5134.
Abstract 5136: HER2 or EGFR inhibition interacts with tumor microenvironment by downregulation of PD-L1 and chemokines
Authors: Kim Jin Won , Sung Ji Hea , Kang Mi Hyun , Kim Ji-Won , Kim Se-Hyun , Lee Keun-Wook , Lee Hye Seung , Kim Jee Hyun
Published: July 2016

Background:

Agents targeting HER2 or EGFR have been applied in several cancer types. Tumor microenvironment including tumor-infiltrating lymphocytes could be a predictive marker in HER2- or EGFR- targeted therapy, although there are some controversies. However, the effects of HER2 and EGFR inhibition on tumor microenvironment are unclear.

Method:

We screened HER2, EGFR, and PD-L1 expression in gastric cancer cell lines by western blot (SNU216, SNU668, SNU719, AGS, N87, YCC3, and YCC10). HER2 and EGFR was inhibited by using dual inhibitor (afatinib, lapatinib). After HER2 and EGFR inhibition, the change of PD-L1 expression was evaluated in HER2 overexpressed cell lines (SNU216, N87, SKBR3) at western blot, FACS, PCR, and qPCR. Selective blockade for down-steam molecules of HER2 and EGFR was conducted by using pictilisib (PI3K inhibitor) and trametinib (MEK inhibitor). The change of chemokines such as CXCL1, CCL2, and CCL21 was evaluated after HER2 and EGFR inhibition by PCR. In clinical data, PD-L1 expression and HER2 expression in resected gastric cancer were also evaluated by immunohistochemistry analysis.

Results:

EGFR-overexpressed or HER2-overexpressed gastric cancer cell lines (SNU216, SNU668, N87) showed higher protein expression of PD-L1. PD-L1 expression decreased with dose-dependent manner in afatinib- or lapatinib- treated cell lines (SNU216, N87, SKBR3). In pictilisib-treated cell lines, PD-L1 was also down-regulated. However, trametinib-treated cell lines did not show down-regulation of PD-L1. After lapatinib treatment in HER2 overexpressed cell lines, CXCL1, CCL21, and CCL2 decreased with dose-dependent manner. In 289 patients with resected gastric cancer, there was a significant association between HER2 and PD-L1 expression (p = 0.03).

Conclusions:

PD-L1 is associated with HER2 and EGFR expression. PD-L1 is regulated by inhibition of PI3K pathway. Therefore, HER2 or EGFR inhibition could interact with tumor microenvironment by down-regulation of expression of PD-L1 and chemokines.

Citation Format: Jin Won Kim, Ji Hea Sung, Mi Hyun Kang, Ji-Won Kim, Se-Hyun Kim, Keun-Wook Lee, Hye Seung Lee, Jee Hyun Kim. HER2 or EGFR inhibition interacts with tumor microenvironment by downregulation of PD-L1 and chemokines. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5136.
Abstract 514: Effect of radiation therapy on breast epithelial cells in BRCA1/2 mutation carriers
Authors: Chiang Huai-Chin , Elledge Richard , Larson Paula , Jatoi Ismail , Li Rong , Hu Yanfen
Published: July 2016

Women carrying BRCA1 and BRCA2 mutations have significantly elevated risk of developing breast and ovarian cancers. BRCA1-associated breast cancer likely originates from progenitors of the luminal epithelial lineage. Recent studies indicate that radiation therapy (RT) for BRCA1 cancer patients is associated with lower incidence of developing subsequent ipsilateral breast cancer. In the current study, we analyzed tumor-free breast tissue procured via prophylactic bilateral mastectomy from three BRCA1 and one BRCA2 mutation carriers, who had been previously treated with RT for unilateral breast cancers. Freshly isolated breast cells from the irradiated and non-irradiated breasts of the same individuals were subjected to flow cytometry, using established cell-surface markers. Two out of the three BRCA1 carriers and the BRCA2 carrier exhibited significantly diminished luminal cell population in the irradiated breast versus the non-irradiated side. The remaining BRCA1 carrier showed similar cell abundance between the two sides. Our finding suggests that prior RT could result in preferential depletion of the luminal epithelial compartment and thus reduced incidence of BRCA1-associated breast cancer. Further investigation may inform development of RT-based preventive approaches aimed at specifically decimating the source of BRCA1-associated tumors.

Citation Format: Huai-Chin Chiang, Richard Elledge, Paula Larson, Ismail Jatoi, Rong Li, Yanfen Hu. Effect of radiation therapy on breast epithelial cells in BRCA1/2 mutation carriers. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 514.
Abstract 5142: Correlation of peripheral blood S100A9+ myeloid-derived suppressor cells and tumor-associated macrophages in lung adenocarcinoma patient
Authors: Lee Kang-Yun , Feng Po-Hao
Published: July 2016

Background: Peripheral blood S100A9+ monocytic myeloid derived suppressor cells (MDSCs) is a predictive factor for treatment response of chemotherapy and epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI) of lung adenocarcinoma patients. Tumor associated macrophages (TAMs), which might derived from peripheral blood monocyte, is also a predictive factor to treatment response of EGFR-TKI. However, the correlation of S100A9+ monocytic MDSCs and TAMs is not well understood in lung adenocarcinoma patients.

Methods: Stage IV lung adenocarcinoma patients with paired formalin fixed paraffin embedded (FFPE) tumor tissue and peripheral blood mononuclear cell (PBMCs) were enrolled. CD68+ cells were calculated in 3 different 200X microscopic field of FFPE as TAMs, and S100A9+ cells also calculated with same method as MDSCs in tumor microenvironment. S100A9+ MDSCs percentage was calculated by flow cytometry from CD14+S100A9+ in PBMC.

Results: Twenty-four lung adenocarcinoma patients with paired FFPE tissue and PBMC data were enrolled. PBMC S100A9+ MDSCs percentage is 17.9± 13.6; average number of S100A9+ cells and CD68+ TAMs in tumor tissue is 96.8±45.7 and 79.3±13.3, respectively. The correlation between S100A9+ MDSCs percentage and tissue S100A9+ cells and CD68+ TAMs is significant (Spearman r = 0.54, p = 0.003 for S100A9+; Spearman r = 0.03, p = 0.03 for CD68; n = 24) CD68+ and S100A9+ cells were also correlated (Spearman r = 0.41, p = 0.04, n = 24). Clinically, high or low (cutoff value as median number) tissue S100A9+ cells and CD68+ TAMs were both predictive factors for treatment PFS for EGFR-TKI. (For CD68, median: 56, high vs low TAM PFS = 7.4 vs 15.4 months, p = 0.04; For S100A9+ cells, median: 76, high vs low S100A9+ cells PFS = 7.2 vs 13.6 months, p = 0.05)

Conclusion: Peripheral blood S100A9+ MDSCs is well correlated to tumor tissue S100A9+ cells and CD68+ TAMs. Both tissue S100A9+ cells and CD68+ TAMs were predictive factors for EGFR-TKI PFS. Our study suggests that peripheral blood S100A9+ monocytic MDSC might be one of the origin of tumor tissue CD68+ TAMs.

Citation Format: Kang-Yun Lee, Po-Hao Feng. Correlation of peripheral blood S100A9+ myeloid-derived suppressor cells and tumor-associated macrophages in lung adenocarcinoma patient. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5142.
Abstract 5145: To explore the tumorigenic regulation mechanism of SPZ1 in hepatocellular carcinoma
Authors: WANG LI-TING , HSU SHIH-HSIEN
Published: July 2016

Cancer directly affects at least one-third of the human population. Despite this extensively, the genetic determinants of cancer risk remain unknown. Spermatogenic leucine zipper 1 (SPZ1), a basic helix loop helix leucine zipper (bHLH-Zip) transcription factor, acted as a proto-oncogene in mouse and human. It has been demonstrated that SPZ1 was involved in early embryonic development, cell growth and differentiation. In this study, we compared the expression of SPZ1 in 168 paired specimens of HCCs and adjacent normal tissues and also in paired specimens from 140 patients with non-HCCs. In pathologic analysis, SPZ1 exhibited a tumor-specific expression pattern and a high correlation to patient survival time, tumor size, tumor number, and progression stage. Enforced expression of SPZ1 accelerated cell proliferation and tumorigenic activity in hepatoma cells. In contrast, short hairpin RNA-mediated attenuation of SPZ1 in hepatoma cells decreased cell proliferation and malignant transformation in vitro and in vivo. Together, we speculated that SPZ1 as a potential therapeutic target of HCCs

Citation Format: LI-TING WANG, SHIH-HSIEN HSU. To explore the tumorigenic regulation mechanism of SPZ1 in hepatocellular carcinoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5145.
Abstract 5150: Identification of higher mRNA expression of both Kitenin and ErbB4 CYT-2 isoform as a molecular marker for the prediction of transition from colon adenoma to carcinoma
Authors: Bae Jeong A , Sun Eun Gene , Ko Yoo-Seung , Choi Hui Jeong , Bae Woo-Kyun , Cho Sang-Hee , Chung Ik Joo , Moon Kyung-Sub , Yu Young-Hyun , Ha Hyung-Ho , Kim Hangun , Kim Kyung Keun
Published: July 2016

Purpose: The molecular events in the malignant progression of colon adenoma after loss of adenomatous polyposis coli (APC) are not fully understood. KITENIN (KAI1 C-terminal interacting tetraspanin) increases the invasiveness of colorectal cancer (CRC) cells and we recently identified a novel EGFR-independent oncogenic signal of EGF that works under co-expressed KITENIN and ErbB4. Here we tested whether elevated KITENIN and ErbB4 contributes to further progression of intestinal adenoma following APC loss.

Results & Discussion: The intestinal tissues of villin-KITENIN transgenic mice in which villin-driven elevated KITENIN induced increased c-Jun expression exhibit mild epithelial cell proliferation but no epithelial lineage changes, compared with those of non-transgenic mice. Among the 4 ErbB4 isoforms, JM-a/CYT-2 and JM-b/CYT-2 exhibited the highest AP-1 activity when cells co-expressing KITENIN and each isoform were stimulated by EGF. Interestingly, predominant overexpression of ErbB4 CYT-2 isoform mRNA as well as increased EGFR expression were observed in intestinal adenoma of APCmin/+ mice making the microenvironment of activated EGF signaling. When we crossed villin-KITENIN mice with APCmin/+ mice, intestinal tumor tissues in the crossed mice showed the characteristics of early stage invading adenocarcinoma. In CRC patients, ErbB4-CYT-2 mRNA expression was significantly greater in tumor tissues than in normal adjacent tissues, but no differences in tumoral expression between the stages, and a positive correlation exists between KITENIN and ErbB4-CYT-2 mRNA expression. Thus, elevated expression of KITENIN and ErbB4-CYT-2 is newly identified as a factor that promotes the transition of colon adenoma to adenocarcinoma within an APC-loss-associated tumor microenvironment.

Conclusion: Higher mRNA expression of both KITENIN and ErbB4-CYT-2 in colon adenoma tissues can be used as a molecular marker of the possible development of malignant transformation.

Citation Format: Jeong A Bae, Eun Gene Sun, Yoo-Seung Ko, Hui Jeong Choi, Woo-Kyun Bae, Sang-Hee Cho, Ik Joo Chung, Kyung-Sub Moon, Young-Hyun Yu, Hyung-Ho Ha, Hangun Kim, Kyung Keun Kim. Identification of higher mRNA expression of both Kitenin and ErbB4 CYT-2 isoform as a molecular marker for the prediction of transition from colon adenoma to carcinoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5150.
Abstract 5152: Inhibition of TLR/IRAK pathway in hepatocellular carcinoma augmented therapeutic responses
Authors: Cheng Yik Ling Bowie , Ng Oi Lin Irene , Lee. Kin Wah Terence
Published: July 2016

Hepatocellular carcinoma (HCC) is the second most lethal cancer worldwide. Frequent relapse and drug resistance accounts for the poor prognosis, which could be attributed by the presence of cancer stem cells (CSCs), a subpopulation that harbor the properties of self-renewal, clonal tumor initiation and long-term repopulation capacity. Toll like receptors (TLRs) are widely expressed on immune cells and TLR/Interleukin-1 receptor associated kinase (IRAK) pathway was originally best known for their role in the induction of innate immune responses, but it was recently found to be frequently up-regulated in various cancers, although its role in cancer development remains largely unexplored.

By RNA sequencing analysis of 16 pairs of HCC clinical samples, we found IRAK1 to be significantly overexpressed in HCC among all members of IRAK family. By quantitative PCR and western blot analyses, overexpression of IRAK1 at both mRNA and protein level was confirmed, which correlated with advanced tumor stage (p = 0.04). By lentiviral-based over-expression and knockdown approaches, we demonstrated that IRAK1 promoted cell proliferation and invasiveness. Interestingly, IRAK1 regulated CSC properties including tumorigenicity, self-renewal, drug resistance and expression of liver CSC markers including CD24 and CD47. To further examine the therapeutic potentials of targeting IRAK1 in HCC, we treated HCC cells with IRAK1/4 inhibitor. Suppression of IRAK1 at 10μM not only consistently inhibited CSC properties, but also augmented the effect of sorafenib, a molecularly targeted drug for treatment of advanced HCC. The cancer-promoting and drug desensitization effect of IRAK1 could be partly explained by activation of NF-κB signaling pathway as well as down-regulation of apoptotic cascade.

Conclusion:

Overexpression of IRAK1 in HCC promotes cancer progression by enhancing proliferation and invasiveness; Importantly, IRAK1 may promote cancer stemness and confers resistance to sorafenib. All in all, targeting IRAK1 with specific small molecule inhibitor alone or in combination with other drugs may be a novel therapeutic regimen for treatment of HCC.

Citation Format: Yik Ling Bowie Cheng, Oi Lin Irene Ng, Kin Wah Terence Lee. Inhibition of TLR/IRAK pathway in hepatocellular carcinoma augmented therapeutic responses. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5152.
Abstract 5158: The change of EZH2 expression in development of colorectal cancer from adenoma
Authors: Ohuchi Mayuko , Nakamoto Yasuo , Tokunaga Ryuma , Nakamura Kenichi , Kosumi Keisuke , Harada Kazuto , Shigaki Hironobu , Kurashige Junji , Iwatsuki Masaaki , Baba Yoshifumi , Miyamoto Yuji , Yoshida Naoya , Baba Hideo
Published: July 2016

[Background]Colorectal cancer has been revealed to develop from adenoma through mutation and deletion of various genes such as FAP, KRAS, DCC and p53 in the process called adenoma-carcinoma sequence. Furthermore, proteins of the polycomb repressive complex 2 (PRC2) function as transcriptional repressors by trimethylating histone H3 at lysine 27, and this complex's activity is essential for cell proliferation and differentiation. The histone methyltransferase enhancer zeste homolog 2 (EZH2), a key member of PRC2, is associated with transcriptional repression of tumor suppressor gene. In our previous study, EZH2 expression was reported to increase as pathology worsened in pancreatic IPMN. Thus EZH2 expression was hypothesized to increase in the adenoma-carcinoma sequence as well.[Purpose]The purpose of this study was to investigate the change of EZH2 expression in progressing colorectal cancer. [Materials and Methods]63 patients with colorectal adenoma, 24 patients with carcinoma in adenoma and 14 patients with colorectal carcinoma who underwent surgical or endoscopic resection were enrolled in this study. The normal lesions of the clinical specimens were also examined. We evaluated the association between EZH2 expression, pathology and expression of tumor suppressor gene. Immunohistochemistry of EZH2, Ki-67, p21, p27 and p16 was performed.[Results]The Ki-67 expression increased parallel to the worsening of pathological findings (p = 0.009), and EZH2 expression showed a significantly positive association with Ki-67 expression (p = 0.02). There was a significant increase of EZH2 expression between normal and colorectal adenoma and carcinoma in adenoma. However, no significance between carcinoma in adenoma and colorectal cancer in EZH2 expression could be identified. Conversely, p21 expression decreased significantly between normal and adenoma lesion; however, p16 and p27 expression showed no major change. [Conclusion]EZH2 expression considerately increased parallel to pathological worsening and p21 expression showed a decrease between normal and colorectal adenoma. Thus EZH2 can contribute to development of colorectal cancer from adenoma via p21 suppression.

Citation Format: Mayuko Ohuchi, Yasuo Nakamoto, Ryuma Tokunaga, Kenichi Nakamura, Keisuke Kosumi, Kazuto Harada, Hironobu Shigaki, Junji Kurashige, Masaaki Iwatsuki, Yoshifumi Baba, Yuji Miyamoto, Naoya Yoshida, Hideo Baba. The change of EZH2 expression in development of colorectal cancer from adenoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5158.
Abstract 5159: Smad7 knockdown in colon cancer cells activates protein kinase RNA-associated eIF2α pathway thereby leading to cell death
Authors: De Simone Veronica , Bevivino Gerolamo , Sedda Silvia , Izzo Roberta , Fantini Massimo Claudio , Monteleone Giovanni
Published: July 2016

Background. Up-regulation of Smad7, an inhibitor of TGF-β1, occurs in sporadic colorectal cancer (CRC). Knockdown of Smad7 with a specific antisense oligonucleotide (AS) leads to activation of eIF2α, an attenuator of protein synthesis, and arrest of CRC cells in the S phase of the cell cycle with the downstream effect of inducing cell death.

Aim. To investigate the mechanisms by which Smad7 knockdown activates eIF2α.

Methods. Phosphorylation of eIF2α was evaluated in CRC cell lines (i.e. HCT116 and DLD-1) either untreated or treated with Smad7 sense (S) or AS by Western blotting (WB) and immunofluorescence (IF). Expression of ATF4 and CHOP, two downstream targets of EIF2α, were evaluated by IF and activation of PKR, GCN2 and PERK, up-stream kinases that induce eIF2α phosphorylation, was assessed by WB. Wild type or PKR-deficient CRC cells treated with Smad7 AS were monitored for eIF2α activation and induction of death. Finally, we assessed whether enhanced phosphorylation of eIF2α seen in cells treated with Smad7 AS was also associated with reduced interaction between eIF2α and PP1, a phosphatase that normally dephosphorylates eIF2α.

Results. Smad7 knockdown increased ATF4 and CHOP expression thus confirming previous data showing activation of eIF2α phosphorylation in CRC cells treated with Smad7 AS. Among kinases that induce eIF2α phosphorylation, only PKR was activated by Smad7 knockdown. Consistently, silencing of PKR reduced but did not abolish Smad7 AS-induced eIF2α phosphorylation and cell death, thus suggesting the existence of further mechanisms that control eIF2α phosphorylation in Smad7-deficient cells. Indeed, in CRC cells, Smad7 interacted with PP1 and Smad7 knockdown reduced association of PP1 with eIF2α.

Conclusions

Data show that Smad7 is involved in CRC cell survival and suggest that Smad7 is a valid target for therapeutic intervention in CRC.

Citation Format: Veronica De Simone, Gerolamo Bevivino, Silvia Sedda, Roberta Izzo, Massimo Claudio Fantini, Giovanni Monteleone. Smad7 knockdown in colon cancer cells activates protein kinase RNA-associated eIF2α pathway thereby leading to cell death. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5159.
Abstract 516: The mechanism associated with radio-resistant renal cell carcinoma and its targeting strategy
Authors: Yun Eun-Jin , Lin Chun-Jung , Saha Debabrata , Hsieh Jer-Tsong
Published: July 2016

Introduction and Purpose: Although ionizing radiation (IR) represents an effective regimen for most solid tumors, renal cell carcinoma (RCC) is known to be highly radio-resistant and the underlying mechanisms associated with IR-resistance remained elusive. In this study, we unveiled the factors associated with IR-resistance and the potential targeting strategy.

Experimental procedures: Several RCC sublines were generated with different genetic manipulation and cells were irradiated with different doses and the surviving fractions were evaluated in 7 days. To investigate the underlying mechanism, various biochemical and molecular biologic methods, such as quantitative real-time PCR, mass-spectroscopy (MS), immunoprecipitation (IP), western blot, and ELISA were performed.

Results: DAB2IP (DOC-2/DAB2 interactive protein) known as a tumor suppressor that functions as a scaffold protein modulating many signal pathways associated cell growth/survival, apoptosis/autophage, and cell migration/invasiveness. In RCC, the status of DAB2IP has not been characterized. We found DAB2IP was frequently lost in RCC; more than 50% in every subtype of RCC. We observed that DAB2IP-deficient RCC cells exhibited IR-resistance whereas re-introduction of DAB2IP could sensitize RCC cells to IR. From MS and IP, it appeared that DAB2IP could directly interact with poly (ADP-ribose) polymerase-1 (PARP-1). In the presence of DAB2IP, PARP-1 activities were suppressed because DAB2IP was able to induce PARP-1 protein degradation via unique proteasome pathway. Since PARP-1 is commonly involved in many cellular functions such as DNA damage repair or transcription modulation, which is also considered as a therapeutic target particularly IR therapy. In this study, elevated PARP-1 led to rapid DNA repairing and IR resistance in DAB2IP-expressing RCC cells. In contrast, by knocking-down PARP-1 in DAB2IP-deficient RCC cells can sensitize cells to IR, suggesting that PARP-1 inhibitors can be a potential radio-sensitizer.

Conclusion: In summary, this study demonstrates a new functional role of DAB2IP in IR-resistant RCC cells via modulating PARP-1 expression and activities. Therefore, targeting PARP-1 by small molecular inhibitor or gene therapy becomes a potential therapeutic strategy to overcome IR resistant RCC.

Citation Format: Eun-Jin Yun, Chun-Jung Lin, Debabrata Saha, Jer-Tsong Hsieh. The mechanism associated with radio-resistant renal cell carcinoma and its targeting strategy. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 516.
Abstract 5161: Development of cutaneous basal cell carcinomas from cutaneous actinic keratoses
Authors: Lambert W Clark , Gagna Claude E. , Lambert Muriel W.
Published: July 2016

Actinic keratoses (AKs) are extremely common proliferations of epidermal keratinocytes which show cytological atypia and impaired ability to mature as the cells that comprise them migrate from the basal layer of the epidermis to higher levels of the epidermis. A small percentage of AKs are well known to progress to cutaneous squamous cell carcinomas (SCCs). However, progression of AKs to cutaneous basal cell carcinomas (BCCs) has not been convincingly documented despite several attempts, even though BCCs are significantly more common than SCCs in the general US and European populations. Documented cases have been dismissed as actinic keratoses that have simply occurred superimposed upon basal cell carcinomas due to chance occurrence of two common sun induced skin lesions at the same sites.

In a university based referral dermatopathology service, one of us (WCL) observed 21 cases of AKs progressing to BCCs during a 29 year period (January 1, 1985 to December 31, 2013) of continuous observation in which 115,898 cases of AKs, 10,188 cases of BCCs and 4,809 cases of SCCs were diagnosed. Only cases in which a continuous progression of cells from those showing changes typical of AKs to those showing changes typical of BCCs was observed were included. During the same interval 308 cases of AKs progressing to SCCs were diagnosed. To further test the hypothesis that AKs can progress to BCCs, cases of AKs that progressed to BCCs were further examined to determine those in which neither the AK nor the BCC occupied the entire skin surface (i.e., length of the epidermal basement membrane) present in the specimen. Of the 21 cases, 13 met this criterion. All were shave biopsies of sun damaged skin. For each biopsy, the proportion of the surface covered by the AK, the proportion occupied by the BCC, and the proportion of the surface overlapped by both lesions were measured and the proportion of overlap was compared to the proportion of overlap expected due to chance. Statistical analysis showed that the observed overlap exceeded the expected overlap by 32 per cent with p < 0.05. We conclude that progression of AKs to BCCs, while not as common as progression of AKs to SCCs, does occur in a small proportion of cases.

Citation Format: W Clark Lambert, Claude E. Gagna, Muriel W. Lambert. Development of cutaneous basal cell carcinomas from cutaneous actinic keratoses. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5161.
Abstract 5162: Id1 is an oncogenic target of RNA binding protein HuR in gastric cancer
Authors: Han Shuang , Wu Xiaoqing , Guo Yuxiao , Zhang Jing , Gu Xuan , Marquez Rebecca T. , Aube Jeffrey , Xu Liang
Published: July 2016

Hu antigen R (HuR), a ubiquitous RNA-binding protein (RBP), has been shown to be overexpressed in gastric cancer. Cytoplasmic HuR is reported to be associated with high stage, distant metastases and unresectability. HuR efficiently regulates cancer-related gene expression by binding to AU- and U-rich sequences (ARE) in the 3’ and 5’-untranslated region (UTR) of mRNAs. Id1 (Inhibitor of DNA binding) is a putative oncogene that we previously found was upregulated in gastric cancer and associated with poor prognosis. Id1 mRNA contains several putative HuR ARE sites. Our goal is to determine the impact of HuR on Id1 expression. We first confirmed Id1 is a HuR target by Ribonucleoprotein Immunoprecipitation (RNP-IP) in gastric cancer cells. Then, we found KH-3, a potent and specific inhibitor of HuR, inhibited Id1 mRNA and protein expression. The proliferative and invasive capabilities of the gastric cancer cells were also inhibited by KH-3. We conclude that Id1 is a target of RNA-binding protein HuR in gastric cancer. HuR specific inhibitor, KH-3, may be a promising therapeutic agent in treating HuR/Id1 high gastric cancers.

Citation Format: Shuang Han, Xiaoqing Wu, Yuxiao Guo, Jing Zhang, Xuan Gu, Rebecca T. Marquez, Jeffrey Aube, Liang Xu. Id1 is an oncogenic target of RNA binding protein HuR in gastric cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5162.
Abstract 5176: Orthotopic syngeneic tumor models for preclinical evaluation of cancer immunotherapy strategies
Authors: Zhang Juan , Guo Sheng , Wang Zhensheng , Li zhongliang , Qiao Meng , Shi Qian
Published: July 2016

Background: The recent successes of cancer immunotherapies have stimulated interest in the potential application of these approaches in many solid and hematologic tumors. Syngeneic tumor models have been widely used in cancer immunotherapy assessment. However, most of these experimental models were established as subcutaneous models, while there may be significant difference in immune microenvironment comparing orthotopic vs. subcutaneous tumors. In addition, drug delivery and metabolism are different in orthotopic tumors vs. subcutaneous tumors. To address this unmet need, we set out to validate a panel of orthotopic syngeneic models for anti-cancer therapeutics (mainly immunotherapeutics) evaluation.

Results: A large collection of orthotopic syngeneic models covering breast cancer, liver cancer and various hematological cancers were established in immunocompetent mice. Survival curves were established and the responsiveness to anti-PD1, anti-PDL1 and anti-CTLA4 antibodies were evaluated in the orthotopic models. Orthotopic liver tumors were monitored with ultrasound imaging to obtain longitudinal growth curves. The correlation in drug efficacy between the orthotopic model and its subcutaneous counterpart was analyzed. The role of NK cells in the development of a panel of systemic hematological tumor models was investigated with NK depleting antibodies.

Conclusion: Despite the obvious difficulties, orthotopic syngeneic models represent closer model systems to human diseases and may be a valuable tool to the evaluation of cancer immunotherapies and their combination strategies.

Citation Format: Juan Zhang, Sheng Guo, Zhensheng Wang, zhongliang Li, Meng Qiao, Qian Shi. Orthotopic syngeneic tumor models for preclinical evaluation of cancer immunotherapy strategies. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5176.
Abstract 5177: RNAseq and FACS profiling of syngeneic mouse models treated with immune checkpoint inhibitors enable biomarker discovery and model selection for cancer immunotherapy
Authors: Zhang Lan , Zhang Juan , Guo Sheng , Qian Wubin , Wang Zhensheng , Shi Qian
Published: July 2016

Background: Syngeneic tumor models have long been used in cancer research. Recently, the clinical success of anti-CTLA4 and anti-PD1 antibodies resulted in increased interests in using syngeneic models to evaluate cancer immunotherapeutics. Furthermore, as researchers discovered chemo, radio and targeted therapies of cancer may interact or change tumor immune environment, they are looking for suitable models that may evaluate the combination of those therapies. More importantly, in the clinic it is still unknown why some patients respond to certain immunotherapies while others do not. We set out to utilize syngenetic models to address those questions.

Material and methods: Syngeneic cell line models were used to evaluate anti-PD1, PD-L1 and CTLA4 antibodies efficacies. 13 tumors from different syngeneic models were collected before the treatment for RNAseq analysis. Tumors were also collected after treatment for immune cell analysis with FACS for PD response. Biomarkers were analyzed with the RNAseq data to predict treatment response to different immune checkpoint inhibitors.

Results: Crown has established the largest collection of syngeneic models with well characterized immunotherapy data. Those models have a diverse response to anti-PD1, PD-L1 and CTLA4 antibodies, ranging from close to 100% inhibition to inducing tumor growth upon antibody treatment. Mostly recently, we have generated detailed maps of the expressional and mutational profiles of those models. Mutational analysis indicated a number of syngeneic models harbor mutations that may be useful for combination studies of targeted and immuno-therapy. Chemo and targeted therapy in combination with immunotherapy in syngeneic models suggested potential strategies that may be successful in the clinic. Analysis of RNAseq and FACS analysis data indicated markers that may be useful to predict immunotherapy response.

Conclusions: These data will enable selection of models for chemo or targeted therapies in combination with immunotherapy. In addition, predictive biomarkers obtained from the analysis may be useful in understanding patient response in the clinic.

Citation Format: Lan Zhang, Juan Zhang, Sheng Guo, Wubin Qian, Zhensheng Wang, Qian Shi. RNAseq and FACS profiling of syngeneic mouse models treated with immune checkpoint inhibitors enable biomarker discovery and model selection for cancer immunotherapy. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5177.
Abstract 5179: Development of a humanized mouse model for direct evaluation of anti-human PD-L1 antibodies
Authors: Qiao Meng , Zhang Juan , Ding Jian , Zhang Rui , Li Zhongliang , Zheng Jia , Lu Teng , Ning Jinjing , Shi Qian
Published: July 2016

Therapies that perturb the binding of programmed death-1 receptor ligand (PD-L1) to its receptor PD-1 achieved unprecedented rates of sustained clinical responses in patients with various types of cancer. Mouse surrogate antibodies were initially evaluated in syngeneic mouse models as the proof of concept for anti-PD-L1 therapy.

However, there's an urgent need to develop appropriate animal model to directly evaluate anti-human PD-L1 antibodies before clinical trial. Here we describe our ongoing efforts in developing a chimeric mouse/human cell line to test human anti PD-L1 antibody. We profiled PD-L1 expression by FACS using anti-mouse PD-L1 antibody and confirmed expression of PD-L1 in a series of murine cancer cell lines. H22, a liver cancer line with moderate expression level of PD-L1 was selected due to its in vivo sensitivity towards anti-PD1 and anti-PD-L1 antibodies. Crispr/cas9 system was employed to knockout murine PD-L1 and replace it with the human counterpart. The targeted knockout had been confirmed by sequencing, the expression of human PD-L1 is illustrated by FACS using antibody that specifically recognize human form and no cross reaction to mouse form. The engineered H22-hPD-L1 cell line was then inoculated subcutaneously to immune-competent BALB/c mice to establish the in vivo model that maintains complete murine immune system and harbor humanized PD-L1. The tumor growth curve of H22-hPD-L1 model and its sensitivity to anti-human PD-L1 antibody have been established.

In summary, our H22-hPD-L1 model may be a valuable tool to evaluate the in vivo activity of anti-human PD-L1 antibody therapies either as a single agent or combinational strategies. The similar engineering may be applied to more murine cell lines (e.g. EMT-6, B16F10 and MBT-2, etc) to provide a spectrum of cell lines with different diseases and genetic makeup for immunotherapies involving anti-hPD-L1 antibodies.

Citation Format: Meng Qiao, Juan Zhang, Jian Ding, Rui Zhang, Zhongliang Li, Jia Zheng, Teng Lu, Jinjing Ning, Qian Shi. Development of a humanized mouse model for direct evaluation of anti-human PD-L1 antibodies. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5179.
Abstract 518: Ionizing radiation-induced DNA damage response decreases during induced pluripotent and embryonic stem cell differentiation
Authors: Mujoo Kalpana , Pandita Raj K. , Tiwari Anjana , Charaka Vijay , Hittelman Walter N. , Hegde Murlidhar , Wang Haibo , Hunt Clayton R. , Dave Bhuvanesh , Chang Jenny , Butler E Brian , Pandita Tej K.
Published: July 2016

Embryonic as well as induced pluripotent stem cells (ESC/iPSC) have a robust DNA damage response to facilitate efficient repair of DNA strand breaks as well as interstrand cross-links (ICLs). Stem cell abundance in tumors can impact the efficacy of the therapeutic agents, and such agents also have an impact on the normal tissue toxicity. Therefore, it is important to compare DNA damage repair in progenitor stem cells and their differentiated lineages since the normal tissues surrounding the tumor predominantly contain differentiated cells. We therefore asked whether the DNA damage response is altered during differentiation of embryonic as well as induced pluripotent stem cells and found that sensing of ionizing radiations, interstrand cross-linking or hydroxyurea induced DNA damage was minimally altered by cellular differentiation. Furthermore, DNA DSB repair by non-homologous end joining was minimally impacted. However, the homologous recombination of DSB repair or ICL repair was significantly reduced in differentiated cells. In addition, differentiated cells showed a reduction in the frequency of new origins of replication and increased stalled replication forks. Interestingly, similar to ESC or iPSC lines, breast tumor stem cells also exhibited a decrease in DNA damage repair by homologous recombination in differentiated cells. The defect in DNA damage repair by homologous recombination was reversed in de-differentiated cells, supporting the argument that differentiation can influence homologous recombination repair of DSBs as well as repair of ICLs. These studies have implications in modulating the therapeutic efficacy of DNA damage inducing tumor-killing agents.

Citation Format: Kalpana Mujoo, Raj K. Pandita, Anjana Tiwari, Vijay Charaka, Walter N. Hittelman, Murlidhar Hegde, Haibo Wang, Clayton R. Hunt, Bhuvanesh Dave, Jenny Chang, E Brian Butler, Tej K. Pandita. Ionizing radiation-induced DNA damage response decreases during induced pluripotent and embryonic stem cell differentiation. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 518.
Abstract 5180: Bayesian multi-endpoint analysis of syngeneic mouse immunotherapeutic trials
Authors: Gold David , Chen Ningning
Published: July 2016

Recent and notable immunotherapeutic successes for the treatments of melanoma and lung cancer have catalyzed wide-spread development of novel immunotherapies and combinations for a variety of cancers. While promising, responses to immunotherapies are a focal point of present investigations, being as of yet challenging to explain. Amongst the many translational challenges, pre-clinical in bred syngeneic mouse experiments often display heterogeneous tumor responses, whereby a fraction of tumors show resistance, partial, or complete response to the same immuno-intervention(s) implanted in identical in bred mouse strains.

We mathematically modeled heterogeneous tumor responses in syngeneic mouse trials with Bayesian statistical methodology. Our decision criteria for drugs and drug combinations can simultaneously evaluate two or more endpoints. At the heart of our methodology is Bayesian learning with prior information that did not compromise statistical performance; actually, we achieved statistical gains to identify superior immunotherapies, rendering quantifiable efficiencies by design. We illustrate an augmented approach to discovery by Bayesian learning in real syngeneic mouse studies. We propose our powerful multi-endpoint Bayesian statistical criteria to ascertain superior pre-clinical immunotherapeutic activity.

Citation Format: David Gold, Ningning Chen. Bayesian multi-endpoint analysis of syngeneic mouse immunotherapeutic trials. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5180.
Abstract 5185: Development of patient-derived xenograft (PDX) models for pancreatic carcinoma as a preclinical platform for drug development
Authors: Ricono Jill , Thatte Jayant , Scott Colleen , Broudy Thomas B.
Published: July 2016

Pancreatic ductal adenocarcinoma (PDA) remains one of the most aggressive tumors in humans. A notable feature of PDA is its innate resistance to many chemotherapies. In preclinical studies, Abraxane showed antitumor activity as a single agent and synergistic activity in combination with gemcitabine in murine models of pancreatic cancer. In this study, we have developed and characterized multiple models of patient derived xenograft (PDX) pancreatic models. These models recapitulate major structural and genetic features noted in patient populations. Developed PDX models are from both prior treated and non prior treated patients with varying levels of resistance to Gemcitibine. Here we have investigated efficacy of standard of care agents in these pancreatic models and demonstrate the potential utility of the pancreatic models in drug discovery efforts in oncology for the treatment of pancreatic cancer.

Citation Format: Jill Ricono, Jayant Thatte, Colleen Scott, Thomas B. Broudy. Development of patient-derived xenograft (PDX) models for pancreatic carcinoma as a preclinical platform for drug development. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5185.