bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 121: Identification of tumor suppressive microRNAs in maxillary sinus squamous cell carcinoma based on microRNA expression signature
Authors: Hanazawa Toyoyuki , Nohata Nijiro , Kikkawa Naoko , Mutallip Muradil , Katayama Akihiro , Harabuchi Yasuaki , Okamoto Yoshitaka , Seki Naohiko
Published: June 2012

Maxillary sinus squamous cell carcinoma (MSSCC) comprises 2-3% of all cancers of head and neck tumor and the annual incidence rate is 0.5-1.0 per 100,000 populations. After introduction of multimodality treatment comprising radiation therapy along with concomitant intra-arterial chemotherapy, rates of local control was improved. However the prognosis of advanced MSSCC is still worse. The 5-years survival rate of T4 tumors is 50% around. Local recurrence is the most common cause of treatment failure and death. Greater understanding of the molecular oncogenic pathways in MSSCC could help improve diagnosis, therapy, and prevention of the disease.

MicroRNAs (miRNAs), non-coding RNA 21-25 nucleotides in length, regulate gene expression primarily at the posttranscriptional level. Growing evidence suggests that miRNAs are aberrantly expressed in many human cancers, and that they play significant roles in carcinogenesis and cancer progression. The expression signature of cancer related miRNAs could contribute as useful biomarkers for diagnosis and improvement of prognosis for MSSCC.

No studies have been carried out for the purpose of identifying specific miRNA alterations in MSSCC. In this study, we performed the miRNA expression signature specific to MSSCC by evaluating 749 mature miRNAs expressions in 5 matched pairs of MSSCC tissues and normal maxillary sinus epithelium. A subset of 9 miRNAs was significantly down-regulated in our signature. Gain-of-function analysis revealed that miR-874 was significantly inhibited cell proliferation, suggesting miR-874 has a tumor suppressive function in MSSCC. We are continuing a search of target genes of miR-874 by using genome-wide gene expression analysis.

The identification of tumor suppressive miRNAs and their target genes could provide new insights into potential mechanism of MSSCC carcinogenesis.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 121. doi:10.1158/1538-7445.AM2011-121
Abstract 1216: The Fn14 receptor is highly expressed in NSCLC tumors and in NSCLC cell lines harboring EGFR-activating mutations
Authors: Tran Nhan L. , Asrani Kaushal , Jameson Nathan , Inge Landon , Cheng Emily , Hostetter Galen , Weiss Glen , Tuncali Serdar , Bremner Ross , Winkles Jeffrey A.
Published: June 2012

Lung cancer is the leading cause of cancer deaths in the USA and worldwide and ∼85% of these cancers are of the non-small cell lung cancer (NSCLC) subtype. Approximately 10-15% of the NSCLC patients in the USA and 30-50% of these patients in Asia have tumors harboring somatic mutations in the epidermal growth factor receptor (EGFR) that cause constitutive activation of this receptor. These patients have the best clinical response to the small molecule EGFR tyrosine kinase inhibitors (TKIs) gefitinib and erlotinib. Here we report that the TNF receptor superfamily member fibroblast growth factor-inducible 14 (Fn14) is frequently overexpressed in NSCLC tumors. We have also found that NSCLC cell lines that contain the same EGFR activating mutations found in patients express high levels of Fn14. Erlotinib treatment of these cells decreases Fn14 levels; therefore, EGFR signaling is indeed triggering Fn14 gene expression in these cell lines. Consistent with this proposal, when EGFR activation mutant receptors (L858R, ΔL747-E749, and D770-N771 insertion) were expressed in rat RL65 lung epithelial cells Fn14 expression was induced. In contrast, RL65 cells expressing a kinase-deficient EGFR receptor (D837A) did not show an increase in Fn14 expression. Since some NSCLC tumors with EGFR activating mutations acquire an additional EGFR mutation (T790M) that promotes TKI resistance, we examined Fn14 levels in a cell line containing both types of mutations. The Fn14 receptor was expressed at high levels in these cells. Finally, we found that shRNA-mediated Fn14 knockdown reduces NSCLC cell migration. Together, these data indicate that EGFR activation in NSCLC cells increases Fn14 gene expression and that Fn14 levels remain elevated in drug resistant cells. Furthermore, Fn14 may play a role in NSCLC cell motility. We propose that Fn14 may be a good biomarker for selection of NSCLC patients most likely to benefit from EGFR TKIs. It may also be a therapeutic target for NSCLC patients; in particular, for those patients with EGFR-driven tumors who have either primary or acquired resistance to EGFR tyrosine kinase inhibitors.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1216. doi:10.1158/1538-7445.AM2011-1216
Abstract 122: Downregulated miR-375 in tongue cancer and its putative role as a tumor suppressor microRNA
Authors: Jung Hyun Min , Patel Rushi S. , Cheng Jin Q. , Chan Edward K.L.
Published: June 2012

MicroRNAs are approximately 22-nt RNAs involved in posttranscriptional regulation of target gene expression. Oral cancers are ranked as the sixth most prevalent cancer worldwide, with squamous cell carcinoma accounting for the most common type (83% – 90%). Evolution and progression of oral cancers is thought to result from the accumulation of genetic alterations in molecular pathways. In order to identify aberrantly expressed miRNAs in oral squamous cell carcinomas (OSCCs), miRNA microarray was performed using 3 healthy controls and 17 OSCC tissue samples all derived from human tongue. Clustering expression patterns of miRNAs in the 17 OSCC samples using Pearson correlation coefficient demonstrated TNM stage-dependent miRNA expression pattern in OSCC. In the expression profile, 134 miRNAs (105 upregulated miRNAs and 29 downregulated miRNAs) were identified to be aberrantly expressed in OSCCs (p < 0.05). The expression of miRNAs was validated by quantitative RT-PCR. MiR-375, the most underexpressed miRNA, was 7.2-fold (p=0.004) downregulated in OSCC samples compared to 5 healthy tongue samples. When compared to the control tongue samples, 88% (15/17) of OSCC samples showed lower miR-375 levels. Furthermore, the expression of miR-375 was inversely proportional to the TNM stage score (stage 2: 32.7-fold, p=0.008; stage 4: 38.4-fold, p=0.002). MTS assay showed that transfected miR-375 mimic inhibited cell proliferation of human oral cancer cell line CAL 27 in dose dependent manner. YWHAZ (14-3-3ζ), PDK1, JAK2, and ELAVL4 (HuD) were reported as miR-375 targets in other cancers. These 4 putative miR-375 targets were examined in CAL 27 cells. JAK2 and ELAVL4 were barely detectable in CAL 27 and thus not significantly regulated by miR-375. In contrast, YWHAZ and PDK1 were repressed both at the mRNA (54% and 19%, respectively) and protein levels (∼30% for both) after transfection of miR-375 mimic into CAL 27 cells. Since both YWHAZ and PDK1 genes are known to play important roles in proliferation and possibly in apoptosis, miR-375 may be the gatekeeper for the balance of the two cellular processes. The absence of miR-375 expression and its inhibitory role in oral cancer cells implicates miR-375 as a tumor suppressor miRNA.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 122. doi:10.1158/1538-7445.AM2011-122
Abstract 1220: Enigma blocks Cbl-c-mediated RET ubiquitination and degradation
Authors: Kales Stephen C. , Lipkowitz Stanley
Published: June 2012

RET (rearranged during transfection) is an RTK that is activated by glial cell-derived neurotrophic factors (GDNF) and is critical to neuronal and kidney development. RET mutations causing loss of function are associated with Hirschsprung's disease, while gain of function mutations are associated with thyroid carcinomas and mutiple endocrine neoplasias (MEN2A&B). Cbl proteins are a family of ubiquitin protein ligases that mediate RTK degradation and thereby serve as negative regulators of RTK signaling. Cbl-c (a.k.a. Cbl-3), the most recently identified of the human Cbl proteins, is restricted to epithelial cells and most resembles the Cbl proteins found in lower organisms. Cbl-c can ubiquitinate and downregulate RTKs including activated RET and EGFR. Through yeast two-hybrid screening, we identified the LIM protein Enigma (a.k.a. PDLIM7) as a Cbl-c interacting protein. Enigma is an adapter protein for kinase anchoring to the cytoskeleton and is required for the mitogenic signaling of the RET/PTC2 oncogene. The interaction was confirmed in mammalian cells through co-immunoprecipitation of transfected and endogenous Cbl-c and Enigma. Enigma did not co-immunoprecipitate with c-Cbl or Cbl-b indicating that this interaction is specific to Cbl-c. Cbl-c can ubiquitinate and downregulate an activated form of RET, RET-MEN2A and we show that Enigma prevents Cbl-c-mediated RET ubiquitination and downregulation. This effect is specific to RET as EGFR ubiquitination was unaffected by the presence of Enigma. An Enigma mutant, lacking the RET binding site, does not block Cbl-c recruitment and subsequent ubiquitination and downregulation of RET. Our data suggest that Enigma blocks Cbl-c binding to the RET receptor and prevents its ubiquitination. This is further supported by microscopy data which demonstrates maintained surface expression of RET in the presence of Enigma. Together these data demonstrate a novel interaction between a positive and negative regulator or RET signaling.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1220. doi:10.1158/1538-7445.AM2011-1220
Abstract 1225: α-Catulin promotes tumor growth by preventing cellular senescence
Authors: Chen Yuh-Ling , Fan Li-Ching , Liang Chen-Hsien , Tsai Yao-Tsung , Hsu Keng-Fu , Hong Tse-Ming
Published: June 2012

Cellular senescence is a proven tumor-suppressive strategy that forces transformed cancer cells to re-enter irreversible cell-cycle arrest. Triggering the senescence of tumor cells may contribute to successful cancer therapy. α-Catulin, an α-catenin-related protein, is reported to have tumorigenic potential because it regulates the NF-κB pathway, but little is known about its clinical relevance and the mechanism through which it regulates cancer progression. Here, we provide evidence that the expression of α-catulin is upregulated in most human cancer cells and clinical oral cancer tissues and positively correlated with tumor size (P = 0.001) and cancer stage (P = 0.004) of OSCC. α-Catulin knockdown in the OC2 and A549 cancer cell lines dramatically decreased cell proliferation and contributed to cellular senescence, and inhibited OC2 xenograft growth. Mechanistic dissection showed that α-catulin depletion strongly induced the DNA damage response (DDR) via a p53/p21-dependent pathway in A549 cells, but a p53/p21-independent pathway in OC2 cells carrying mutant p53. Global gene expression analysis revealed that α-catulin knockdown altered several crucial genes related to mitotic chromosome condensation, DDR, and DNA repair systems, which suggests that its depletion-induced cellular senescence might be caused by chromosome condensation failures, severe DNA damage, and impaired DNA repair ability. Our study provides evidence that α-catulin promotes tumor growth by preventing cellular senescence and suggests that downregulating α-catulin may be a promising therapeutic approach for cancer treatment.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1225. doi:10.1158/1538-7445.AM2011-1225
Abstract 1226: p53 signaling pathway acts as rescue mechanism to β1 integrin silencing in head and neck cancer cell line
Authors: Kim Mi Ra , Chang Hyo Won , Kim Hyo Jung , Moon So Young , Lee Myung Jin , Han MyungWoul , Kim Seong Who , Kim Sang Yoon
Published: June 2012

β1 integrin's aberrant expression has implicated a cancer progression and resistance to cytotoxic therapy. Moreover, cancer cells gained epithelial mesenchymal transition (EMT) features were more resistant to the detachment-induced cell death. In this regard, we investigated the mechanism by which β1-Integrin governs cell viability and verified whether the signaling pathway is associated with EMT features.

Cancer cell lines from head and neck cancer patients (AMC-HN3 and AMC-HN9) and well-known EMT cancer, MDA-MB231, were used in this study. To determine whether three cell lines reveal the epithelial EMT features, EMT markers such as Snail, vimentin, fibronectin and E-cadherin were evaluated by western blot analysis and immunofluroscence. To knockdown β1 integrin signal, β1 integrin small interfering RNA (siRNA) was transiently transfected into all cell lines. The cell viability was examined by looking at the results of propidium iodide staining, MTT assay, changes in the mitochondrial membrane potential and cell morphology.

MDA-MB231 and AMC-HN9 cells possessed EMT features whereas AMC-HN3 cells were not shown EMT phenotype. β1 integrin silencing cell increased the sub G1 from 1.0% to 29.8%, increased the percentage of cells losing their mitochondrial membrane potential from 5.3% to 65.9% as well as the number of fragmented mitochondria in AMC-HN3 cells. In contrast, MDA-MB231 cells showed more resistance to silencing of β1 integrin, lower cell death rate from 0.6% to 9.6% and less changes in the mitochondrial membrane potential from 4.34% to 34.51%. Interestingly, AMC-HN9 cells showed complete resistance to β1 integrin blocking even though FAK phosphorylation signaling was blocked in AMC-HN9 cells, same as AMC-HN3 cells, which is a different response from typical EMT featured cells. When AMC-HN9 cells were treated to inhibit β1 integrin and p53 separately, it showed no response to changes in the cellular morphology, viability and apoptosis-related signal pathway; however, when it was given the combined β1 integrin and p53, apoptosis occurred in the cell line.

In this study, acquisition of the EMT features in cancer cells presented resistance to the β1 integrin inhibition. In addition, the activation of p53-p21 signaling pathway provided resistance to detachment-induced apoptosis in head and neck cancer cell line with EMT features.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1226. doi:10.1158/1538-7445.AM2011-1226
Abstract 1227: Anti-oncogenic role of Peroxisome proliferator-activated receptor- β/Δ (Pparβ/Δ) involves regulation of viral Hras1 (v-Hras1)-induced senescence and endoplasmic reticulum (ER) stress in mouse primary keratinocytes
Authors: Zhu Bokai , Khozoie Combiz , Bility Moses , Blazanin Nicholas , Glick Adam B. , Gonzalez Frank J. , Peters Jeffrey M.
Published: June 2012

Pparβ/Δ-null mice exhibit enhanced chemically-induced skin tumorigenesis compared to wild-type counterparts. Since an oncogenic mutation of Harvey sarcoma ras virus gene (v-Hras1) is a critical event in chemical carcinogenesis, the role of PPARβ/Δ in mouse primary keratinocytes infected with a retrovirus encoding viral Hras1 (v-Hras1) gene was examined. While v-Hras1 induced senescence in wild-type mouse primary keratinocytes, the level of v-Hras1-induced senescence was significantly reduced in Pparβ/Δ-null keratinocytes, which was due in part to decreased phospho-MEK and phospho-ERK during the early stage of v-Hras1 expression. In addition, while total v-Hras1 protein level was comparable between wild-type and Pparβ/Δ-null keratinocytes, the level of the GTP-bound form of v-Hras1 was significantly decreased in Pparβ/Δ-null keratinocytes from 4 day to 7 day post v-Hras1 infection. Surprisingly, the decreased activity of v-Hras1 did not lead to a decreased level of phospho-AKT in Pparβ/Δ-null keratinocytes. However, Pparβ/Δ-null keratinocytes were more dependent upon PI3K-AKT pathway for maintenance of proliferation and evasion of senescence. Differentially expressed genes between wild-type and Pparβ/Δ-null keratinocytes that are involved in the regulation of senescence were determined by cDNA microarray analysis. Differential expressions of both positive and negative regulators of RAS pathway were observed between wild-type and Pparβ/Δ-null keratinocytes following v-Hras1 infection. Notably, the mRNA encoding CDK inhibitor p27Kip1 was lower in Pparβ/Δ-null keratinocytes, In addition, Pparβ/Δ-null keratinocytes showed enhanced endoplasmic reticulum (ER) stress and unfolded protein response (UPR) post v-Hras1 infection, characterized by enhanced vacuolation and higher levels of major ER stress markers. However, the enhanced ER stress did not lead to apoptosis in Pparβ/Δ-null keratinocytes. These results suggest an anti-oncogenic role of PPARβ/Δ in v-Hras1-transformed mouse primary keratinocytes that involves both the promotion of anti-tumorigenic senescence and attenuation of pro-tumorigenic ER stress and UPR. (Supported by CA124533)

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1227. doi:10.1158/1538-7445.AM2011-1227
Abstract 1230: Wnt5a-dependent induction of senescence suppresses epithelial ovarian cancer
Authors: Bitler Benjamin G. , Nicodemus Jasmine P. , Li Hua , Cai Qi , Soring Keri , Birrer Michael J. , Connolly Denise C. , Godwin Andrew K. , Cairns Paul , Wu Hong , Zhang Rugang
Published: June 2012

Epithelial Ovarian Cancer (EOC) remains the most lethal gynecological malignancy in US and is the fifth leading cause of cancer deaths among American women. Thus, there is an urgent need to understand the etiology of EOC to develop novel therapies for this disease. Wnt5a is classified as a non-canonical Wnt ligand. Here, we demonstrated that Wnt5a is expressed at significantly lower levels in human EOC cell lines and in primary human EOC compared with either normal ovarian surface epithelial cells (p=0.039) or fallopian tube epithelial cells (p<0.001). Importantly, expression of Wnt5a in primary human EOC inversely correlates with tumor stage (p=0.003) but not tumor grade (p=0.086). Interestingly, Wnt5a expression is significantly lower in Type II high-grade serous EOC compared to Type I EOC that includes low-grade serous, mucinous, clear cell and endometrioid subtypes of EOC (p=0.005). In addition, we discovered that hypermethylation of promoter CpG island contributes to Wnt5a downregulation in human EOC cells. Significantly, restoration of Wnt5a expression in human EOC cells promoted senescence of EOC cells and resulted in a dramatic decrease in cell proliferation both in vitro and in vivo in an orthotopic model of EOC in the ovary of SCID mice. Mechanistically, Wnt5a inhibited canonical Wnt/beta-catenin signaling and resulted in the activation of senescence-promoting histone repressor A/PML pathway. In summary, we show that Wnt5a is often expressed at lower levels in primary human EOC and Wnt5a expression suppresses the growth of EOC cells by triggering senescence through antagonizing canonical Wnt signaling. Our data imply that the cell of origin between Type I and Type II EOC is different. These results also suggest that loss of Wnt5a expression is a putative marker of EOC and that non-canonical Wnt signaling is a potent target for developing novel EOC therapeutics.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1230. doi:10.1158/1538-7445.AM2011-1230
Abstract 1232: The loss of p16INK4a function is essential for Werner Syndrome mouse embryo fibroblasts to escape from senescence
Authors: Zhang Xiufeng , Wu Xiaomin , Tang Wenru , Luo Ying
Published: June 2012

p16INK4a is a tumor suppressor gene contributing to cell cycle arrest as a cyclin dependent kinase inhibitor. It has been shown that the expression of p16INK4a protein is up-regulated along with the cellular senescence and the loss of p16INK4a function could elongate the cellular life span. These data suggest that p16INK4a promote cellular senescence. However, it is still debated that the expression regulation of p16INK4a in mouse cells is due to cell culture condition.

Previous study has showed that mouse embryo fibroblasts (MEFs) from late generation of Werner Syndrome (WS) mice (G5 mTR−/− Wrn−/−) senesced very rapidly, and the senescent MEF cells are prone to escape senescence and spontaneously immortalize.

To understand the molecular basis for WS MEFs escaping from senescence, we compared the expression level of proteins involved in regulating cellular senescence. Western blotting showed that all immortalized WS MEFs lost their expression of p16INK4a and re-expression of p16INK4a could induced cellular senescence, suggested the loss of p16INK4a function is essential for maintaining senescence status of WS MEFs.

To further evaluate the role of p16INK4a in senescence induced by telomere dysfunction, p16INK4a-/− mice were crossed with Wrn−/− mice to generate MEFs with genotypes of p16INK4a-/−, Wrn−/− and p16INK4a-/−Wrn−/−. Growth curve of these three cell lines revealed that Wrn−/− MEFs grew slower than p16INK4a-/− MEFs, the p16INK4a-/−Wrn−/− MEFs grew about the same as p16INK4a-/− MEFs. This data suggested that p16INK4a-/− rescued the growth barriers occurred in Wrn−/− MEFs. In consistent with this, Western blotting showed that loss of p16INK4a in Wrn−/− MEFs (p16INK4a-/−Wrn−/−) could partially eliminate the DNA damage responses caused by Wrn−/−, e.g., the expression of H2AX, p19ARF. Furthermore, telomere dysfunction was initiated by introducing TRF2ΔBΔM into these cells. In response to this stimulation, Wrn−/− MEFs senesced in several passages. In contrast to this, p16INK4a-/− and p16INK4a-/−Wrn−/− MEFs could continue grow and “kicked out” the exogenous TRF2ΔBΔM in their late passages. In summary, our data suggested that in the context of telomere dysfunction, loss of p16INK4a function could prevent the cells from senescence.

This work was supported by The New Century Excellent Scholarship to Y.L, NSFC grant to Y.L. (30771194, 30970598) and to W.T. (30960152). We thank Dr. Sandy Chang from Yale University and Dr. Ronald A Depinho from Harvard University for providing transgenic mice used in this study.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1232. doi:10.1158/1538-7445.AM2011-1232
Abstract 1234: Development of a quick sensitive real-time PCR assay for measuring telomere length
Authors: Qin James , Calado Rodrigo
Published: June 2012

Telomeres are the ends of linear chromosomes with tandem hexameric nucleotide repeats. Its function is to protect the natural ends of chromosomes from being recognized as damaged DNA, hence contributing to chromosomal stability. Telomeric sequences shorten each time the DNA replicates; when the telomere shrinks to a certain threshold, the cell can no longer divide. Its metabolism slows down, the cell ages, and dies. This molecular clock mechanism is also associated with cancer progression and therapy. Turning off telomerase activity by shutting off telomerase activation in cancer cells will led to cellular senescence. The current gold standard method to measuring telomere length is by Southern blot; however, it is tedious and costly, and often takes up to two days to complete. Here, we describe a new, simplified automated real-time PCR assay to accurately detect the abundance of these 5’-TTAGGG-3’nucleotide repeats and determine frequency and the length of Telomere in 47 min.

The assay demonstrated to be highly reproducible and accurate for human peripheral cells.

The result has good correlation of R2=0.9939, slope=3.41, sensitivity to 1ng of gDNA and has an amplification efficiency of 100%. We evaluated this assay with 299 healthy human subjects to date ranging from 0 (core blood) to 99-year of age. The results are positive with an inverse correlation between telomere length and age, as telomeres were shorter with aging. The telomere length was measured in 13 samples using the gold standard Southern blot method and the new q-PCR method (R2 = 0.8623), significantly higher than the originally described (Cawthon NAS 2002).

We believe this fast, sensitive methodology is a powerful tool and provides a new avenue for investigators to address the full extent and significance of telomere shortening in the tumorigenic process. This new assay described here will allow for more widespread use of this technique among cancer researchers and for study of age related telomere diseases and conditions.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1234. doi:10.1158/1538-7445.AM2011-1234
Abstract 1237: Role of Src in resistance to anoikis in detachedpancreatic cancer cells
Authors: Gomes Evan , Connelly Sarah F. , Summy Justin M.
Published: June 2012

Pancreatic adenocarcinoma is an aggressive malignancy currently ranked as the fourth leading cause of cancer related death in the United States, with over 95% of patients succumbing to the disease within 5 years of diagnosis. The vast majority of pancreatic cancer deaths are due to metastatic dissemination of the primary tumor. The non-receptor protein tyrosine kinase, c-Src (Src) has emerged in recent years as a potential target for pancreatic cancer treatment. In vitro studies have demonstrated the importance of Src for pancreatic tumor cell adhesion, migration, invasiveness, and resistance to apoptotic death. Mouse models of pancreatic cancer have indicated an important role for Src in formation of metastases. In addition, SHP-2, a non-receptor protein tyrosine phosphatase that regulates Src family kinase activity, has been recently shown to be a key mediator of various signaling pathways and evidence has established clinical relevance for SHP-2 in human diseases. Our goal is to determine the importance of Src for discrete steps in the progression to pancreatic cancer metastasis and resistance to detachment-induced cell death and the role of SHP-2 in modulating Src activity and downstream biology during this process. We have demonstrated a rapid and robust activation of Src in L3.6 human pancreatic cancer cells that have become detached from the extracellular environment, a crucial step in the development of metastases. Also, we have observed co-immunoprecipitation between Src and SHP-2 in detached cells, suggesting a potential role for SHP-2 in regulating Src activity in detached pancreatic cancer cells. In addition, our results reveal a Src-dependent activation of the Akt cell survival cascade and the stress kinase, Jun N-terminal kinase (JNK) in detached cells. Furthermore, this study confirmed that inhibition of both Src and SHP-2 in combination sensitizes L3.6 pancreatic cancer cells to anoikis in detached cells. We hypothesize that Src activity in detached pancreatic cancer cells promotes metastasis through suppression of anoikis. We will confirm these in vitro studies by testing the effects of inhibition of Src alone or in combination with SHP-2 inhibition on metastatic tumor formation using a nude mouse experimental metastasis model. The potential use for the Src and SHP-2 signaling axes as therapeutic targets for many cancers remains largely untapped. Our studies suggest that the anti-tumor and anti-metastatic effects of Src inhibition may be improved through simultaneous inhibition of SHP-2.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1237. doi:10.1158/1538-7445.AM2011-1237
Abstract 1243: Cofilin mediates actin destabilization and shape change in replicative senescence and stress-induced premature senescence
Authors: Tsai Cheng-Han , Lin Liang-Ting , Chou Yen-Ting , Lee Yi-Jang
Published: June 2012

Cellular senescence is an irreversible growth arrest mechanism that prevents indefinite proliferation of mammalian cells. Change of morphology is a common characteristic of senescent cells, while the underlying mechanism has not been completely addressed. In this study, we found that the flattened and enlarged cell shape in aged WI-38 fibroblast was accompanied by reduced actin cytoskeletal organization and up-regulated cofilin-1, a member of actin depolymerization factor family. Up-regulation of cofilin-1 was also detected in various tissues (lung, kidney, heart and liver) of one-year old BALB/c mice compared to that of 6-week old mice. Forced expression of cofilin-1 in early passaged cells accelerated senescence, while knockdown of cofilin-1 in late passaged cells inhibited senescence. Hydrogen peroxide mediated stress-induced premature senescence (SIPS) also induced cofilin-1 in human lung cancer cell lines, and this phenomenon was suppressed by knockdown of cofilin-1. Up-regulation of cofilin-1 in SIPS was independent of p53, while p27kip1 was essential for cofilin-1 induced senescence. Knockdown of p27kip1 in cells treated with hydrogen peroxide or in that with over-expressed cofilin alleviated senescence. Moreover, forced expression of cofilin-1 was sufficient to induce senescence in human lung cancer cells and repress the growth of xenograft tumor in NOD-SCID mice. Taken together, we found that cofilin-1 is able to mediate the cell senescence that is important for repressing the cancer development.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1243. doi:10.1158/1538-7445.AM2011-1243
Abstract 1245: Characterization of human leukemic cell lines and changes of the characteristics according to passage number
Authors: Hur Eun-Hye , Kim Sung-Doo , Goo Bon-Kwan , Lee Sung-Sook , Lee Kyoo-Hyung , Lee Jung-Hee , Kim Dae-Young , Choi Yunsuk , Lee Je-Hwan
Published: June 2012

Background: Human leukemic cell lines are valuable tools for the studies on the biology of hematologic cancers and new drug development. We investigated the baseline characteristics of human leukemic cell lines and changes of the properties according to passage number.

Materials and methods: A total of 11 human leukemic cell lines were investigated in this study, including myeloid leukemia cell lines (HL60, NB4, Molm-13, MV4-11, KG-1, K562) and lymphoid leukemia cell lines (CCRFCEM, SR, U937, Molt4, Jurkat). Genetic mutations (N-Ras, K-Ras, B-Raf, FLT3-ITD, FLT3-TKD, MLL) and polymorphisms (MDR3435CT and GST genes [A, M, T]) were detected using appropriate methods such as PCR, RFLP or DNA sequencing. MDR activities were measured using Rhodamine 123 efflux assay. Establishment of xenograft model for each cell line was tried with subcutaneous injection of leukemic cells mixed with matrigel matrix into Balb/c nude mouse. Cell lines were maintained with two times of passage per week. Western blot assay was done for changes of expression of intracellular signaling pathway proteins such as ERK, p-ERK, AKT, p-AKT, and HR23B according to the passage number.

Summary of results: N-Ras mutation was found in HL60 (Gln61Leu) and Molt4 (Gly13Val), K-Ras mutation in CCRFCEM (Gly12Asp) and NB4 (Ala18Asp), B-Raf mutation in none, FLT3-ITD and MLL rearrangements in Molm-13 and MV4-11, and FLT3-TKD in none. High MDR activity was noted in KG-1 (92.5%) and other cell lines showed less than 30% of MDR activities. Mouse xenograft model was established with 7 leukemic cell lines (HL60, NB4, MV4-11, Molm-13, K562, Molt4 and CCRFCEM). ERK phosphorylation was observed in most cell lines except CCRFCEM, but AKT phosphorylation was not found in any of the cell lines. As increasing the passage number, the levels of p-ERK in K562 and HL60 significantly decreased whereas those of p-AKT in Molm-13 were elevated.

Conclusion: Our study results provide genetic and biologic data for human leukemic cell lines and show the importance of passage number of the leukemic cell lines in designing experiments.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1245. doi:10.1158/1538-7445.AM2011-1245
Abstract 1249: Metabolomic analysis of a genetically defined model of oncogenesis reveals that common oncogenic mutations result in synergistic deregulation of multiple metabolic pathways
Authors: Schafer Xenia , Smith Bradley , Spencer Cody , Land Hartmut , Munger Joshua C.
Published: June 2012

Oncogenesis induces profound changes to the cellular metabolic network. A number of these cancer-cell metabolic phenotypes have been found to be important for in vivo tumorigenesis. We have utilized liquid chromatography tandem mass spectrometry (LC-MS/MS) to globally profile the metabolic network of a genetically-defined experimental model of oncogenesis in an attempt to identify novel metabolic activities induced by transformation. Furthermore, we sought to delineate how common oncogenic mutations contribute to specific metabolic phenotypes. We have analyzed how dominant negative p53 and constitutively active Ras expression, either independently or in concert, impacts the metabolic activity of non-transformed parental colon crypt epithelial cells. In addition to measuring steady-state metabolite concentrations, we have employed isotope tracer analysis to measure the impact of oncogenic mutation on metabolic pathway kinetics. We find that combined p53 and Ras oncogenic mutation synergistically activates numerous aspects of central carbon metabolism including glycolysis, glutamine catabolism, and pyrimidine and purine biosynthesis. Our data indicate that many well-defined oncogenic metabolic phenotypes are the result of a cooperative effect of combinatorial mutation highlighting the importance of identifying the interaction between oncogenic signal transduction and metabolic regulation for understanding the etiology of the cancer-cell metabolic state.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1249. doi:10.1158/1538-7445.AM2011-1249
Abstract 125: Exogenous MiR-125b inhibits head and neck squamous cell carcinoma growth by suppressing Bcl-2 expression
Authors: Zhao Yuan , Pang Xiaowu , Califano Joseph , Gu Xinbin
Published: June 2012

PURPOSE: Head and neck squamous cell cancer (HNSCC) is one of top ten cancers in the United States. The overall 5-year survival rate of HNSCC has not improved in the past two decades. B-cell lymphoma 2 (Bcl-2), an oncogene, is a key regulator of the apoptosis to promote cell proliferation and apoptosis arrest. Therefore, targeting Bcl-2 is an attractive strategy for restoring a normal apoptotic process and inhibiting cancer cell growth in HNSCC. MicroRNAs (miRNAs) are a class of ∼22-nucleotide-long, endogenously expressed, highly conserved of noncoding RNAs with important regulatory functions in proliferation, apoptosis and metastasis. In this study, we screened eight cancer-associated miRNAs levels (mir-21, -124, -125b, -147, -200a, -200b, -200c and -429) in twenty five human tissue specimens and four cell lines, including twenty HNSCC specimens and five normal controls, and three HNSCC cell lines (JHU-13, -22 and -29) and one transformed oral keratinocytes cell line (OKF-6). Based on the screening data, we further investigated the mechanism of miRNA-125b (miR-125b) regulated Bcl-2 expression and the therapeutic potential of exogenous miR-125b in a miR-125b stable transfected HNSCC cell line. Design Methods: The miRNA expression level was determined by quantitative RT-PCR. A stable expressed exogenous miR-125b HNSCC cell line (JHU-22miR125b) was generated by using lentivirus transfection. Colony formation and MTT assayed cell growth and cell viability and Western blot was used to analyze protein expression. Results: The expression levels of mir-124 and mir-125b were significant lower in both tumor tissue and tumor cell lines than control, and mir-21 was significant higher in tumors. Thus, mir-125b was selected as a candidate for further study. The level of miR-125b in the transfected cell lines (JHU-22miR125b) was increased more than five times compared with the control cells (Transfected with vector only, JHU-22vector). The levels of Bcl-2 protein and cell viability were significantly lower in JHU-22miR125b than JHU-22 vector. CONCLUSION: Bcl-2 is a direct target in miR-125b regulated Bcl-2 expression. Exogenous miR-125b suppresses the proliferation and growth of cultured HNSCC cells. The effect of exogenous miR-125b on inhibition of HNSCC growth via suppressing Bcl-2 expression will be further validated in tumor xanografts. This work was supported in part by grants P20 CA118770 from National Cancer Institute.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 125. doi:10.1158/1538-7445.AM2011-125
Abstract 1261: A preneoplastic switch to glycolytic metabolism driven by a PI3K-induced transcriptional program
Authors: Antico Arciuch Valeria G. , Cristofano Antonio Di
Published: June 2012

Most cancer cells exhibit a peculiar metabolic behavior, producing energy by aerobic glycolysis instead of oxidative phosphorylation (Warburg effect). Several oncogenes directly drive this switch modulating the levels and activity of, among others, Hexokinase II, Lactate Dehydrogenase A, Pyruvate Kinase M2, and Pyruvate Dehydrogenase Kinase 1.

We have generated a Pten-/- mouse model in which the PI3K pathway is selectively activated in the thyroid epithelial cells, leading to hyperplasia which later progresses to follicular carcinoma.

Using proteomics to identify proteins differentially expressed in early hyperplastic lesions, we have found that the levels of many tricarboxylic acid cycle proteins are down-regulated in mutant thyroids. Down-regulation of these proteins is due to reduced mRNA expression, and results in reduced oxidative phosphorylation and enhanced glycolysis (over 3-fold increase in lactate production in mutants vs. wt). Furthermore, although mutant thyroids cells have a normal number of mitochondria, the expression of most mitochondrially-encoded genes is heavily down-regulated. Strikingly, none of the genes and proteins classically associated with the Warburg effect is found deregulated in the mutant thyroid cells.

Analysis of the expression of transcription factors and co-activators involved in cellular metabolism revealed that the decrease in TCA cycle and OXPHOS genes is due to a reduction in ERRα, ERRγ, PPARα and PPARγ levels. Conversely, none of the co-activators analyzed (PRC1, PGC-1α, and PGC-1β) exhibited changes in mRNA expression. Pharmacological and genetic PI3K inhibition in mouse tumor-derived thyroid cancer cell lines increased the expression levels of both TCA cycle OXPHOS genes. Moreover, it also increased mtDNA amount, suggesting that the transition from hyperplasia to cancer involves additional alterations of the metabolic machinery. Likewise, PI3K inhibition induced a marked decrease in lactate secretion. Finally, these thyroid cancer cell lines exhibited high sensitivity to glycolysis inhibitors such as 3-bromopyruvate.

Based on these compelling data, we propose a novel mechanism leading to the Warburg effect in preneoplastic lesions: PI3K activation initiates a coordinated rearrangement of the expression of metabolic genes, which favors aerobic glycolysis at the expense of TCA/OXPHOS. The elucidation of this process will lead to innovative targeted approaches to selectively disrupt tumor progression, while preserving normal metabolism in non mutated cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1261. doi:10.1158/1538-7445.AM2011-1261
Abstract 1264: BRCA1 induces reversion of “Warburg effect” in a breast cancer cell model
Authors: Privat Maud , Morvan Daniel , Guerrab Abderrahim El , Aubel Corinne , Bignon Yves-Jean
Published: June 2012

BRCA1 is a major tumor suppressor in breast cancer and it was implicated in numerous pathways resulting in anticarcinogene functions. Likewise, the hypermetabolic nature of cancer cells and their increased reliance on “aerobic glycolysis”, as originally described by Otto Warburg and colleagues, are considered metabolic hallmarks of cancer cells. The objective of our study was to address specific contributions of BRCA1 to the signature metabolic features of cancer cells, including the so-called “Warburg effect”.

The breast cancer cell line carrying the 185delAG mutation of the BRCA1 gene, SUM1315MO2 cells, was stably transfected with a wild type BRCA1 plasmid. In this cell model, we combined analysis of cell metabolism major enzymes transcriptional expression by TaqMan low density array with the investigation of metabolites by 1H-NMR spectroscopy. We demonstrated that BRCA1 transfection in mutant cells induced strong decrease of glycolysis metabolites and enzymes while TCA cycle and oxidative phosphorylation tended to be activated. Glucose assay confirmed that BRCA1-mutated cells consumed more glucose than cells expressing wild-type BRCA1. Preliminary results seem to show that BRCA1 could act on glycolysis via inhibition of the HIF1α pathway.

Our study showed a new function for BRCA1 in cell metabolic regulation, globally resulting in reversion of the Warburg effect in our breast cancer cell model.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1264. doi:10.1158/1538-7445.AM2011-1264
Abstract 1266: Oncogenic stress induced by Bcr-Abl over-activation leads to cell death mediated by a massively enhanced aerobic glycolysis
Authors: Dengler Michael A. , Staiger Annette , Gutekunst Matthias , Hofmann Ute , Aulitzky Walter E. , van der Kuip Heiko
Published: June 2012

It is well established that normal cells respond to over-activation of oncogenes by induction of genetically encoded programs such as apoptosis or senescence. Therefore, too much activity of a growth promoting oncogene seems to disturb the cellular homeostasis, a phenomenon which is known as oncogenic stress. To investigate the molecular mechanisms behind this phenomenon we used cell clones overexpressing the oncogene Bcr-Abl and cultivated them continuously in the presence of the Bcr-Abl inhibitor Imatinib. Removal of Imatinib led to a robust induction of Bcr-Abl autophosphorylation and concomitant overstimulation of the Bcr-Abl downstream pathways PI3K/AKT-, RAS/MAPK-, and JAK/STAT. Importantly, this acute over-activation of Bcr-Abl resulted in a delayed non-apoptotic cell death starting not before 48 hours after Imatinib withdrawal. This cell death was preceded by a massively enhanced aerobic glycolysis and glutaminolysis (Warburg effect) and amino acid metabolism leading to an elevated cellular ATP and protein content. During the first 30 hours after Imatinib deprivation the enhanced metabolism had no effect on proliferation but resulted in cellular swelling, cytoplasmic vacuolization, and massive induction of ER stress markers indicating that Bcr-Abl over-activation induces ER stress response. However, cell death upon Bcr-Abl over-activation was not dependent on this severe ER stress since siRNA mediated knockdown of CHOP and BIM had no effect on cell viability. In contrast, both inhibition of aerobic glycolysis by 2-deoxy-glucose as well as reduction of glutamine concentration in the medium were sufficient to block Imatinib deprivation induced cell death. These data demonstrate that the Bcr-Abl over-activation mediated enhanced metabolism is responsible for oncogenic stress induced cell death. Interestingly, screening of 1200 marketed small molecule inhibitors (Prestwick chemical library) for their potential effects on oncogenic stress response uncovered selectively steroids to effectively antagonize both ER stress and cell death upon Bcr-Abl over-activation. These agents were also able to normalize cellular metabolism upon Imatinib withdrawal, supporting the view that “overshooting” metabolism upon oncogenic over-activation leads to cell death.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1266. doi:10.1158/1538-7445.AM2011-1266