bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2958: Quantitative and correlative analysis of variant UGT1A mRNA expression in human liver
Authors: Jones Nathan R. , Chen Gang , Lazarus Philip
Published: February 2011

UDP-glucuronosyltransferases (UGTs) play an important role in the metabolism and excretion of various endogenous and xenobiotic compounds including several carcinogens and chemotherapeutic agents. UGTs are divided into two sub-families based on sequence homology. The nine UGT1A family members are encoded by a single locus on chromosome 2q37, where each family member has a unique exon 1 and shares exons 2-5. A recently identified exon 5b in the common region of the UGT1A gene cluster leads to the expression of 18 additional mRNA species from this locus. In vitro studies indicate that the proteins encoded by these variant mRNAs negatively modulate the activity of the wild type isoforms. It is hypothesized that inter-individual variation in the ratio of wild type to variant mRNA expression may affect glucuronidation capacity. The goal of this study was to quantitatively examine variant UGT1A mRNA expression in individual human liver specimens to better assess the importance of splicing variants in overall UGT1A activity. To differentiate between the 5’ (exon 1) and 3’ (exon 5) splice variants, a nested, quantitative PCR reaction was developed that allows both variants to be distinguished simultaneously. Results of the study indicate that there is a 406-, 278-, 58-, and 139-fold inter-individual variation in the expression ratio of wild-type to variant mRNA for UGTs 1A1, 1A4, 1A6, and 1A9, respectively, in the 10 human livers examined. UGT1A variant expression reached as high as 8.2-fold more than wild-type UGT1A1, 2.9-fold higher than wild-type UGT1A4, 0.13-fold lower than wild-type UGT1A6, and 0.46-fold lower than wild-type UGT1A9. Even though the expression ratio of wild-type to variant mRNA differs widely between different individuals, results indicate that the expression ratio is consistent for all UGTs examined within a single individual. The correlation between the expression ratio of UGTs 1A1 and 1A4 in different individuals is highly significant (R2 = 0.9661, p<0.0001). Similarly, the correlation between the expression ratio of UGT1A1 and UGT1A6, UGT1A1 and UGT1A9, UGT1A4 and UGT1A6, UGT1A4 and UGT1A9, and UGT1A6 and UGT1A9 are all significant (R2=0.5265, p=0.0269; R2=0.8994, p=0.0003; R2=0.5782, p=0.0174; R2=0.9610, p<0.0001; R2=0.582, p=0.0277, respectively). Together, these data suggest that UGT1A variant expression could significantly impact overall UGT1A activity and that there is a high correlation between wild-type : variant mRNA expression ratios across the entire UGT1A locus within individuals. Future studies will be conducted to determine whether a difference in the UGT1A expression ratio has an impact on glucuronidation capacity in vivo.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2958.
Abstract 2969: Localization and characterization of ST7 in cancer
Authors: Charong Nurdina , Patmasiriwat Pimpicha , Zenklusen Jean Claude
Published: February 2011

ST7 has been proposed as a tumor suppressor gene at the chromosome region 7q31.1-q31.2. In order to gain some insight into its role in cancer, localization and verification of ST7 expression levels were performed. Various types of ST7 expression vectors tagged with sequences of GFP, YFP or V5 were created using gateway cloning system and full length ST7 cDNA isolated from a human adult brain cDNA library. Cytosolic ST7 expression in HCT-116, MCF-7 and PC-3 cell lines was detected via the fluorescence signal of the fusion proteins. ST7 translocation from cytoplasm to nucleus has not been observed in any of the conditions assayed. A cell cycle synchronization study demonstrated that both ST7 and SERPINE1 were over expressed when cells were arrested. Expression of these genes was found to be substantially diminished when the cells re-entered cell division status. In addition, we also found that Survivin, MMP13, and CyclinD1 were differentially expressed during cell cycle. Our findings suggest that ST7 mediates tumor suppression through regulation of the genes involved in maintaining cellular structure of the cell and involved in oncogenic pathways.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2969.
Abstract 2972: Intestinal p53 deletion leads to accumulation of chromosomal instability without promoting tumor formation
Authors: Begus-Nahrmann Yvonne , Lechel André , Rudolph Lenhard
Published: February 2011

Background:

Colorectal cancer is after lung cancer the second biggest cancer threat for humans and the tumor burden is rising steadily during human ageing. One important issue during human ageing is the accumulation of DNA damage inducing a p53 dependent response leading to p21-dependent senescence or apoptosis. It has been shown that deletion of DNA repair proteins as well as other components of the DNA damage pathway leads to tumor formation.

Methods:

In this approach we investigated the role of the tumor suppressor p53 with and without telomere dysfunction in intestinal tumor formation. Late generation of iG4 mTerc−/−; p53lox/lox and iF1 mTerc+/− p53lox/lox mice were crossed to Villin-Cre-ERT2 transgenic mouse model. In 1 month old mice Tamoxifen was injected on 5 consecutive days leading to functional deletion of the Trp53 gene.

Results:

In aging IF1 Terc+/− p53 deletion did not induce an obvious intestinal phenotype including tumor formation until 24 months. In contrast to p21 deletion the deletion of p53 significantly shortened the lifespan of iG4 Terc−/− Trp53−/− animals compared to iG4 Terc−/− Trp53+/+. This reduction correlated with earlier appearance of crypt atrophy and weight loss but was not associated with tumor formation. Deletion of the Trp53 gene resulted in an abrogation of the p21-dependent senescence and accumulation of DNA damage in the intestinal epithelium of both p53 deleted cohorts. Moreover this correlated with an accumulation of intestinal stem cells carrying high levels of DNA damage. On contrary to p21 deletion the deletion of p53 impaired the depletion of chromosomal instable intestinal stem cells in aging telomere dysfunctional mice.

Conclusion:

These results provide first proof that deletion of p53 is not able to promote tumorigenesis in the intestinal epithelium of mice and that p53 induction has a protective role preventing accumulation of damaged intestinal stem cells in aging telomere dysfunctional mice.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2972.
Abstract 2974: Cep57 is a novel regulator of centriole overduplication in tumor cells
Authors: Cuevas Rolando A. , Duensing Anette , Duensing Stefan
Published: February 2011

Abnormal multipolar mitoses are commonly observed in human cancers where they can increase the risk for chromosome segregation errors, numerical chromosomal instability and malignant progression. The mitotic spindle poles are formed by centrosomes and tumor cells frequently show abnormal centrosomes numbers. The molecular mechanisms leading to numerical centrosome aberrations are not understood in detail. The centrosome consists of two centrioles, which normally duplicate in synchrony with the cell division cycle. We recently identified a novel mechanisms leading to abnormal centriole numbers in tumor cells in which maternal centrioles generate more than the normal one daughter at the same time (centriole multiplication). Using a siRNA library screen enriched for centrosomal proteins, we identified Cep57 as a novel protein involved in centriole multiplication. Cep57 was found to localize to centrosomes and to co-localize and functionally interact with Polo-like kinase 4 (PLK4), a master regulator of centriole duplication control. Cep57 overexpression was found to promote abnormal centrosome and centriole numbers as well as mitotic abnormalities with a significant number of cells showing multipolar metaphases as well as multipolar anaphases and telophases. Cep57 maps to chromosome 11q21, a region that is amplified in several cancer types including hematological malignancies and prostate cancer. Taken together, we identified Cep57 as regulator of centriole duplication in tumor cells and a candidate oncogene. Experiments to target the Cep57-PLK4 axis with small molecules to prevent cell division errors and malignant progression are currently under way.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2974.
Abstract 2980: Role of XPA and p53 genes in the induction of centrosomal amplification in mice exposed to Zidovudine (AZT)
Authors: Nostrand Terri A. , Momot Dariya , Poirier Miriam C. , Olivero Ofelia A.
Published: February 2011

In cultured cells, exposure to the highly-effective nucleoside reverse transcriptase inhibitor (NRTI) Zidovudine (AZT) induces genomic instability, causing cell cycle arrest, micronuclei, sister chromatid exchanges, and shortened telomeres. In previous studies, we demonstrated for the first time that AZT induced centrosomal amplification (defined as > 2 centrosomes /cell), a novel manifestation of genotoxicity. Here, bone marrow cells from wild type C57BL/6J mice and their transgenic counterparts with genotypes XPA −/− p53 +/+, XPA−/−p53 +/− and XPA−/−p53 −/−, were cultured in vitro to obtain homogeneous populations of mesenchymal-derived fibroblasts. Wild type, and DNA repair- / p53-deficient cells (n=3/group) were exposed to 0, 10 or 100 μM AZT for 24 hr, and centrosomal amplification was examined by immunohistochemical (IHC) staining with an anti-pericentrin antibody. Values for centrosomal amplification (% of cells with ≥2 centrosomes/cell) were 12.2, 15.2 and 19.7% in wild type (XPA +/+ p53 +/+) cells exposed to 0, 10 or 100 μM AZT, respectively. Values for centrosomal amplification were 18.0, 28.7 and 33.9 % for XPA −/− p53 +/+cells, and 21.6, 29.3 and 39.5% for XPA−/−p53 +/− cells, after exposure to 0, 10 and 100 μM AZT, respectively. For XPA−/−p53 −/− cells there were 18.4, 22.9 and 29.0% of cells with centrosomal amplification after exposure to 0, 10 and 100 µM AZT, respectively. Statistical analysis (by ANOVA) showed significance between each of the three transgenic groups, when compared to the wild type cells (p= 0.0025). The data suggest that loss of both XPA alleles confers genomic instability, but that additional loss of p53 does not further enhance instability. Whereas nucleotide excision repair is not considered to impact NRTI intracellular processing, these data would argue that lack of intact nucleotide excision repair enhances genomic instability in AZT-exposed cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2980.
Abstract 299: Blockade of Rictor impairs mTORC1 inhibitor-induced upregulation of IGF-IR and Her2 expression in human gastrointestinal cancer cells
Authors: Lang Sven Arke , Moser Christian , Hackl Christina , Fichtner-Feigl Stefan , Schlitt Hans J. , Geissler Edward K. , Stoeltzing Oliver
Published: February 2011

Inhibition of mTOR signaling has recently gained increasing interest for therapy of human gastrointestinal cancer. However, treatment with mTOR inhibitors such as rapamycin only blocks mTORC1 (Raptor, mTOR, mLST8) whereas mTORC2 (Rictor, mTOR, mLST8) is not affected in most cancer cells. This might lead to activation of signaling intermediates such as Akt in mTORC2 dependant manner which has been implicated in resistance to anti-neoplastic therapy. Hence, we hypothesized that inhibition of mTORC2 by Rictor blockade might prevent induction of signaling pathways and improve anti-cancer effects of mTOR inhibitors. Human gastrointestinal cancer cells (TMK-1, HPAF-II) were used for the experiments. mTOR inhibition was performed with rapamycin (Wyeth Phama) and mTORC2 blockade was achieved by siRNA (Invitrogen) against Rictor. Effects on signaling intermediates and receptor expression were determined by Western blotting and RT-PCR, respectively. The results show that mTOR inhibition with rapamycin leads to phosphorylation of Akt on both phosphorylation sites in human cancer cells. mTORC2 blockade by siRNA disrupts activation of Akt as shown by diminished phosphorylation on Thr308. Surprisingly, mTOR blockade with rapamycin showed up-regulation of total IGF-IR and Her2 expression in tumor cells on mRNA and protein level. This effect was reversible by mTORC2 blockade with Rictor siRNA or direct inhibition of Akt signaling using Ly290004. Together, our results show that inhibition of mTOR with rapamycin leads to up-regulation of IGF-IR and Her2 expression in cancer cells in Akt/mTORC2 dependant manner. Blockade of Rictor can prevent these effects providing a basis for combined anti-cancer therapy strategies.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 299.
Abstract 2992: Transcriptional silencing of amplicons by histone deacetylase inhibition
Authors: Kim Tae Hoon , Kim Yoon Jung , Cecchini Katharine R. , Radke Susan , Kumar Amanika , Geyda Kimberly , Narayan Murli , Tuck David , Harris Lyndsay
Published: February 2011

Inhibition of cancer cell growth by histone deacetylase (HDAC) inhibitors has been observed in a number of malignancies. Multiple mechanisms have been proposed to explain the cancer-specific cytotoxic effect of HDAC inhibition, including transcriptional activation of pro-apoptotic and differentiation programs that become epigenetically silenced during tumorigenesis. Here, we report a direct transcriptional repression mechanism of histone deacetylase inhibition that selectively targets genomic amplicons, providing genetic and epigenetic alterations as a basis for the cancer selective cytotoxicity of these drugs. Our study implicates distinct epigenetic structures at the oncogenic amplicons that might be targeted for cancer drug development.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2992.
Abstract 3000: MicroRNA-21 as a biomarker for benefit from adjuvant therapy in resectable pancreatic adenocarcinoma
Authors: Voortman Johannes , Hwang Jin-Hyeok , Giovannetti Elisa , Steinberg Seth M. , Kim Yong-Tae , Funel Niccola , Park Joo Kyung , Kim Min A. , Kang Gyeong Hoon , Kim Sun-Whe , Chiaro Marco Del , Peters Godefridus J. , Giaccone Giuseppe
Published: February 2011

Many pancreatic adenocarcinoma patients who undergo surgical resection receive adjuvant chemotherapy and/or combined modality treatment. However, only a subset of patients benefit from additional therapy. The aim of this study was to evaluate the expression of several potential markers of treatment outcome. Resected tumors of a cohort of 82 pancreatic adenocarcinoma patients from Korea were analyzed for protein expression of 20 markers by immunohistochemistry and expression of 7 microRNAs using quantitative real-time PCR. A second validation cohort of 45 resected Italian patients was analyzed for microRNA-21 (miR-21) expression. Korean and Italian cases consisted of FFPE and frozen specimens, respectively. Univariate and multivariate analyses were used to compare biological parameters and survival endpoints.

In the Korean cohort, Cox proportional hazards model analysis in the subgroup of patients treated with adjuvant therapy (N=52) showed that lower than median miR-21 expression was associated with a significantly lower HR for death (0.316; 95% CI: 0.166-0.600; p-value: 0.0004) and recurrence (0.521; 95% CI: 0.280-0.967; p-value: 0.04). Expression status of miR-21 was the single most predictive biomarker for treatment outcome from all factors evaluated. No significant association of miR-21 expression status with prognosis was detected in patients not treated with adjuvant therapy. In the independent cohort of 45 Italian cases, all treated with adjuvant therapy, lower than median miR-21 expression was also significantly correlated with longer overall and disease-free survival confirming its predictive value.

Concluding, low miR-21 expression was associated with benefit from adjuvant treatment in two independent cohorts of pancreatic cancer cases, using both FFPE and frozen tumor specimens. These data provide evidence that miR-21 expression may allow stratification of pancreatic cancer patients for adjuvant treatment. This is the first strong predictive marker for adjuvant therapy in radically resected pancreatic cancer and further prospective studies are warranted.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3000.
Abstract 3007: Hsa-miR-9 expression patterns are associated with metastatic recurrence in patients with clear cell renal cell carcinoma through hypermethylation of the gene in tumor tissue
Authors: Hildebrandt Michelle A.T. , Gu Jian , Lin Jie , Ye Yuanqing , Tan Weiqi , Tamboli Pheroze , Wood Christopher G. , Wu Xifeng
Published: February 2011

The long-term prognosis for patients with clear cell renal cell carcinoma (ccRCC) is dramatically altered by the development of metastatic recurrence. However, there are very few indicators that can predict which patient will develop a recurrence. MicroRNA gene expression has been shown to be associated with RCC development, and here we show that it can also help to identify patients at increased recurrence risk. We utilized genome-wide microRNA arrays to measure expression levels in tumors from 74 ccRCC patients and identified hsa-miR-9 as a candidate for recurrence risk (Hazard Ratio: 2.6, 95% Confidence Interval: 1.55-4.38) resulting in a reduction in median time to recurrence by 30 months (P-value = 0.0038). This observation was validated in a replication sample of an additional 254 ccRCC tumors. We further showed that the genes encoding for hsa-miR-9 were significantly hypermethylated in tumors compared to adjacent normal tissues (P-value < 0.001 for both miR-9-1 and miR-9-3) resulting in decreased expression, and that the methylation of these genes was more significant in tumor DNA obtained from patients who developed a recurrence (P-value = 0.012 and 0.009 for miR-9-1 and miR-9-3, respectively). Furthermore, methylation of miR-9-1 and miR-9-3 were independently associated with a reduction in time to recurrence. Hsa-miR-9 has been shown to play a role in tumor development for other cancers, and our results demonstrate that it has similar functions in ccRCC while also playing a role in the development of metastatic recurrence.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3007.
Abstract 3011: MicroRNA expression profiles in benign and aggressive canine mast cell tumors
Authors: Fenger Joelle M. , Lin Tzu-yin , Taccioli Cristian , Volinia Stefano , Kisseberth William C. , London Cheryl A.
Published: February 2011

Introduction: MicroRNAs (miRNAs) are small non-coding RNAs (ncRNAs) 18-24 nucleotides in length that regulate gene expression primarily by targeting mRNAs according to the degree of complementarity with their 3’ untranslated region (UTR). MiRNAs are often dysregulated in cancer, suggesting they play a role in tumorigenesis. Canine mast cell tumors (MCT) are a common cutaneous tumor of dogs whose biological behavior varies from relatively benign disease that is cured with surgical excision to aggressive, locally invasive, highly metastatic disease that is poorly responsive even to multimodality therapy. We hypothesize that high grade MCTs possess a unique miRNA expression signature distinct from that found in benign MCTs and that this contributes to their aggressive behavior. In this study, we sought to determine which miRNAs are differentially expressed in benign and aggressive canine MCT.

Methods: Total RNA was isolated by the Trizol (Invitrogen) method from 12 biologically low grade (surgical cure) and 12 biologically high grade (patient died from MCT disease). Mature miRNA expression analysis of preamplified cDNA was done using TaqMan Low Density Arrays (TLDAs), interrogating the expression profile of 378 miRNAs, 151 of whose mature sequences are 100% conserved between human and dog (Sanger miRBase release 12). Normalization was performed with the small nuclear RNA U6 and microRNA expression was calculated utilizing the comparative Ct method. Statistical analysis was performed with RealTime Statminer software (Integromics). P-values of <0.05 were considered statistically significant.

Results: The expression of specific miRNAs was significantly different in biologically low grade and high grade MCTs. 51 miRNAs were differentially expressed in high grade versus low grade tumors (p<0.05). MiRNAs with known importance in human tumorigenesis, including several members of the miR 17-92 cluster (hsa-miR-17, −18a, −18b) and its paralog, the miR-106b-25 cluster (hsa-miR-106b, −93, −25) were significantly overexpressed in high grade tumors as compared to low grade tumors.

Conclusion: Our findings indicate that specific miRNAs contribute to the biologically aggressive canine MCT phenotype. Hierarchical clustering revealed distinct miRNA expression signatures in high grade tumors compared to low grade tumors. Characterization of miRNA expression in canine MCTs will facilitate our understanding the biology of this disease and has the potential to identify diagnostic/prognostic factors and targets for therapeutic intervention.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3011.
Abstract 3012: Development of a microRNA signature for risk of progression in renal cell carcinoma
Authors: Banumathy Gowrishankar , Ibragimova Ilsiya , Al-Saleem Tahseen , Uzzo Robert G. , Cairns Paul
Published: February 2011

The incidence of asymptomatic pathologically localized renal cell carcinoma (RCC) has shown the greatest increase in recent decades. Despite the finding of organ-confined (pT1 or T2) disease at the time of nephrectomy, a subset of patients will have local recurrence or distant metastasis after nephrectomy. Most solid neoplasms are now thought to accumulate a series of genetic and epigenetic changes as they progress through well-defined clinical and histopathological changes. The association of particular molecular markers with progression and outcome of disease means that the status of certain markers can be used to identify the presence of disease and its likelihood for progression. To identify an epigenetic signature that may predict recurrence in patients with organ-confined RCC, we examined RNA hybridized to the Agilent microRNA (miRNA) array from a large number (>100) of RCC patients with extensive clinicopathological annotation. The potential of miRNAs as biomarkers for risk of progression and early detection of recurrent RCC has not yet been studied. Since approximately 90% of metastatic RCC is of clear cell histology, we examined only this histological cell type. The cell type and grade of the RCC was re-reviewed blind by a single pathologist. miRNA expression signatures were established from a series of low grade (I-II), low stage (pT1-2) clear cell RCC with no evidence of disease after nephrectomy for at least 5 years; a series of high grade, high stage (pT3-4), clear cell RCC; and a series of low grade and low stage clear cell RCC that progressed. The expression of particular miRNAs was validated by qRT-PCR. In the growing number of patients who have undergone surgical resection for organ-confined RCC, the near term goal would be to identify those that should be followed more closely and a future goal, depending upon results of ongoing therapeutic trials, to be directed to adjuvant therapy. There would also be cost benefits in potentially following those RCC patients with lowest risk of progression less frequently. MiRNA signatures obtained by our study will address the clinical question whether risk of progression could be defined in patients with organ-confined RCC at initial surgery.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3012.
Abstract 3020: Identification of a subgroup with good prognosis of small-cell lung cancer by miRNA expression profiling
Authors: Oba Takehiko , Ninomiya Hironori , Okumura Sakae , Nishio Makoto , Fujiwara Mutsunori , Ishikawa Yuichi
Published: February 2011

Background: Long-term outcomes of patients suffering from small-cell lung cancer (SCLC), a neuroendocrine carcinoma, are usually poor, but a few patients achieve long-term survival. Prognostic factors in SCLC have not been fully elucidated. Previously we successfully identified a neuroendocrine carcinoma subgroup with good prognosis by mRNA profiling (Jones et al. Lancet 2004). Here we attempted to identify the subgroup by microRNA (miRNA) profiling. miRNAs are known to negatively regulate gene expression and to be relevant to tumorigenesis, tumor classification and prognosis.

Methods 63 samples (46 neuroendocrine tumors including 35 SCLC and 11 large cell neuroendocrine carcinomas (LCNEC), 5 squamous cell carcinoma (Sq), 4 adenocarcinoma and 8 normal lungs) were obtained through surgical resection. miRNA expression profiling was performed using miRNA microarray (Agilent). Hierarchical clustering was performed after selection of about 600 miRNA by choosing those with signal strength more than five.

Results: Unsupervised hierarchical clustering using the 600 miRNA classified the tumors into two subgroups. Group 1 contained 20 SCLC (8 females and 12 males) and 3 LCNEC, and was associated with unfavorable prognosis. Group 2 contained 15 SCLC (1 female and 14 males), 8 LCNEC and 4 Sq, and was with good prognosis. 5-year survival of the two groups was 0% and 79%, respectively (p=0.007). As compared with our former study by mRNA, all 5 cases in the poor prognosis group fell into Group 1, and 6 out of 7 cases in the good prognosis group were included by Group 2, indicating good agreement between mRNA and miRNA profiling. Clinically, tumors of Group 1 tended to be smaller (29mm) than Group 2 (40mm), and cumulative smoking of Group 1 was lower (41 pack-years) than Group 2 (67 p-y) (p=0.03). Serum proGRP levels were significantly higher in Group 1 (50.9 pg/ml) than in Group 2 (26.4) (p<0.01). Among cases treated by preoperative chemotherapy (n=12), PR:NC ratio was 6:3 in Group 1 and 2:1 in Group 2, suggesting no apparent difference of chemosensitivity. A few miRNA were differentially expressed in the two groups.

Conclusion: A distinct subgroup with favorable prognosis existed in SCLC. The subgroup was identified by both miRNA profiling as well as mRNA profiling. Detailed characterization of the subgroup will be presented.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3020.
Abstract 3023: MicroRNA expression profile in pancreatic cancer stem cells
Authors: Jung Dawoon E. , Wen Jing , Oh Tae Yun , Song Si Young
Published: February 2011

MicroRNA(or miRNA) regulates various biological process including development, differentiation, proliferation, maintenance and apoptosis. Recently, existence of cancer stem cells was reported in some solid malignancies, including breast, brain, prostate, liver and pancreas cancers. Pancreatic ductal adenocarcinoma remains an extremely aggressive malignancy that is virtually therapy-resistant. Thus better understanding on cancer stem cell biology will not only help us to develop diagnostic marker for early diagnosis but also to treat against chemoresistant pancreatic cancer. Lately, evidences of miRNA involvement in cancer stem cells have been reported. Several miRNAs including let-7, mir-200, and mir-181 shows regulatory functions in cancer stem cells of different organs, however, miRNAs in other cancers including pancreas has not yet been reported. In this present study, we analyzed pancreatic cancer stem cell specific miRNAs and further investigated their functional role in cancer stem cells.

To analyze the differentially expressed miRNAs, we used sphere cultivation methods to enrich stem cell population and analyzed overall miRNA expression with miRNA microarray. Then correlation of miRNA expression and mRNA expression was analyzed by using linear model Yijkg=µ+Ai+Dj+Vk+Gg+AGig+VGkg+εijkg where A, D, V, and G represent the additive effect of the ith array, jth dye, kth biological group, and gth gene, respectively; AG represents the combinatorial effect of the ith array with the gth gene; εijkg describes the random error associated with each measurement; and VG for the combinatorial effect of the kth biological group with the gth gene. All additive effects were estimated via least squares and an F-test was performed on the modeled data to select differentially expressed genes. The candidate miRNA expression was confirmed by qRT-PCR.

As a result, number of miRNAs showed differential expression in sphere and are considered to have functional role in cancer stem cells by directly or indirectly targeting mRNAs. Further, correlation analysis suggested possible linkage between miRNA clusters with stem cell associated mRNAs.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3023.
Abstract 3026: miR-203 and miR-205 expression profiles are predictive of high grade non-invasive urothelial carcinoma progression
Authors: Rieger-Christ Kimberly M. , Tsai Sheaumei , Lenherr Sara M. , Cimmino Cara B. , Gee Jason R. , Libertino John A. , Summerhayes Ian C.
Published: February 2011

Introduction: MicroRNA (miRNA) has been shown to play a role in regulating the expression of genes involved in tumor suppression and oncogenesis. High-grade, non-muscle-invasive urothelial carcinoma of the bladder (UCB) harbors a significant risk of progression to muscle invasion. In this study, we sought to identify miRNAs differentially expressed in non-progressive high-grade non-muscle-invasive UCB as compared to pathologically identical lesions that progress to muscle invasion.

Methods: Total RNA was extracted from formalin fixed paraffin-embedded tissue (FFPE) of high-grade non-muscle-invasive (TaG3/T1G3) UCB lesions. Patients were categorized into two groups: non-progressors with at least 3 years follow-up or progressors advancing to ≥T2. Exclusion criteria were previous tumors of the upper tract, prior systemic chemotherapy or previous pelvic irradiation. Total RNA was pooled for microarray analysis from a subset of the non-progressor and progressor groups. Data was analyzed using the LOWESS normalization method. Expression profiles of miRNAs were verified with qRT-PCR on 89 high-grade non-muscle-invasive UCB (68 non-progressors and 21 progressors). Logistic regression analysis was performed to identify miRNAs predictive of tumor progression.

Results: MicroRNA microarray data identified a panel of miRNAs (25) with potential involvement in the progression to invasion of high-grade non-muscle-invasive UCB. 36% of these miRNAs were upregulated and 64% were downregulated in the progressor samples. To validate differential expression of these miRNAs, qRT-PCR was performed on the 89 FFPE tissue samples. Logistic regression analysis of qRT-PCR data revealed decreased expression of miR-203 and miR-205 as predictive of progression (p = 0.031 and p = 0.045, respectively). Subsequent qRT-PCR with miR-203 and miR-205 on 31 muscle-invasive tumor samples (≥T2) showed expression profiles that were similar to those of the non-invasive progressors.

Conclusions: In this study, we have shown that miR-203 and miR-205 can identify patients with non-muscle- invasive UCB at high risk to progress to muscle-invasive disease. Our findings also suggest that such tumors have a biomolecular profile similar to muscle-invasive lesions. Further validation studies are underway to investigate additional miRNAs and determine the mechanism by which these miRNAs promote tumor invasion and progression. These results provide an important tool that can be used to prognosticate the clinical behavior of a very challenging subset of tumors.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3026.
Abstract 3035: MicroRNA expression in early stage endometrial cancers
Authors: Chandramouli Gadisetti VR , Shoji Yutaka , Casablanca Yovanni , Maxwell G Larry , Risinger John I.
Published: February 2011

The molecular signatures of endometrial cancers from whole genome gene expression studies are now available setting a stage to explore the key regulators involved in these cancers. Non-coding micro-RNAs (miRNAs) are functionally important regulators of numerous cellular processes affecting growth and dissemination and possibly possess prognostic significance for several cancer types. In a first step toward understanding the miRNA biology of endometrial cancers, we performed an unbiased characterization of about 667 unique human miRNAs in stage-1 endometrial cancers including 51 endometrioid (16 grade-1, 23 grade-2 and 12 grade-3) and 7 serous (grade-3) cancer types and 17 endometrial epithelial samples from healthy individuals. In total, there were 8 stage IA, 32 stage IB and 18 stage IC cancers. RNA was extracted from samples prepared by laser capture micro-dissection which contained minimal non-epithelial cells. TaqMan Low Density Arrays (TLDA) were used to quantitatively determine the expression levels of miRNAs (TaqMan® MicroRNA Arrays A and B from Applied Biosystems). Isolated RNA was transcribed into cDNA and used to quantitatively assess the expression of 768 miRNAs arrayed within the TLDA. The Ct-values were initially normalized by subtracting a reference Ct-value derived from multiple positive controls on the array. These ΔCt values were filtered to exclude the miRNAs that were not expressed in any of the arrays. Statistical calculations were performed on -ΔCt values. In summary, about 158 miRNAs were up or down regulated in cancers at least by four-fold (p<0.001) including many novel candidates for this cancer type. Notably, miR-183* was on average the most differentially over-expressed in both endometrioid and serous cancers compared to normal endometrium. The numbers of miRNAs and their statistical significance to distinguish the histological type, grade, and stage among cancers were found to be in decreasing order respectively. The putative mRNA targets of miRNAs were predicted using Targetscan v. 5.1. These predicted targets were further examined using gene expression data (Affymetrix Human Genome Plus 2.0) of the same RNAs. In conclusion, we identified numerous miRNAs that are disregulated in endometrial cancer. The potential of these miRNAs to define the biology of endometrial cancer is further discussed.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3035.
Abstract 3036: Comparison of miRNA expression between human osteoblasts and osteosarcoma cells
Authors: Duan Zhenfeng , Choy Edwin , Harmon David , Liu Xianzhe , Susa Michiro , Schwab Joe , Mankin Henry , Hornicek Francis
Published: February 2011

Osteosarcoma is the most common primary malignant bone tumor that most frequently affects young adults. There is increasing evidence that miRNAs (miRs) are important modulators of gene expression and their deregulation has been reported in various human tumors. This study investigated differences in miR expression in normal human osteoblast cells versus malignant human osteosarcoma cells differ in their miR expression. Analysis of miRNA expression in normal osteoblast cell lines (hOBc, NHOST, hFOB) and osteosarcoma cell lines (KHOS, Saos, U-2OS) were completed using miR microarray technology with unsupervised hierarchical clustering analysis. The relative expression levels of these miRs were confirmed by real-time quantitative RT-PCR. A number of miRs were found to have different expression in the osteosarcoma cells when compared to normal human osteoblasts. Three miRs, miR-199a-3p, miR-127-3p and miR-376c were significantly decreased in osteosarcoma cell lines while miR-151-3p and miR-191 were increased in osteosarcoma cell lines in comparison to osteoblasts. Because miR-199a-3p has been suggested to play a vital role in lung and liver cancer, we further characterized its biological functional role in osteosarcomas. The precursor (miR-199a-1) of miR-199a-3p was stably transfected into osteosarcoma cell lines KHOS and U-2OS, resulting in a decrease of cellular proliferation. The expression of miR-199a-3p targeted genes in transfected cells was evaluated by Western blot. These targets included MET and mTOR, two genes that were observed to be overexpressed in osteosarcoma tissues and various cell lines. Following miR-199a-3p transfection, a decrease in MET and mTOR expression was observed in KHOS and U-2OS cells. This study demonstrates that various miRs are differentially expressed in osteosarcomas, and miR-199a-3p, in particular may play a functional role in osteosarcoma cell growth and proliferation.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3036.
Abstract 304: Comparison of commercially available phospho-AKT [Ser473] antibodies for immunohistochemical biomarker analysis
Authors: Jeffers Michael , Dauffenbach Lisa M. , Bhargava Ajay , Lathia Chetan , Kerfoot Christopher A.
Published: February 2011

Immunohistochemical (IHC) analysis of AKT phosphorylation on Ser473 is commonly performed on pre- and post-treatment cancer biopsies as part of pharmacodynamic biomarker assessment for targeted oncology therapeutics. The current study was performed to assess the specificity and sensitivity of four pAKT Ser473 monoclonal antibodies for IHC analysis (clones D9E, 736E11, 587F11 and 14-5). To facilitate this analysis, cancer cell lines with constitutive activation of the PI3K pathway due to mutations in PIK3CA (MCF-7 and MKN-1) or PTEN (MDA-MB-468) were grown in the presence or absence of inhibitors of the PI3K pathway (wortmannin and LY294002). Cell lysates were prepared for analysis via Western blotting and formalin-fixed, paraffin-embedded cell pellets were prepared for analysis via IHC. All antibodies produced a band at 65 kDa, the expected molecular weight for pAKT, when used for Western blot analysis of untreated cell lines and demonstrated a reduction or ablation of this band following treatment of the cell lines with inhibitors of the PI3K pathway. When examined via IHC, antibodies 736E11 and 587F11 demonstrated nuclear and cytoplasmic staining in MCF-7 and MKN-1 cells and membrane staining in MDA-MB-468 cells. These antibodies demonstrated a reduction of staining in all three cell lines treated with PI3K inhibitors, but the reduction was modest in MCF-7 and MKN-1 cells. Clone 587F11 was the least sensitive for IHC. Staining with antibodies 14-5 and D9E was predominantly cytoplasmic or membrane associated, while clone14-5 produced a more punctate appearance. Strong staining was achieved with clone 14-5 and prominent reduction or ablation of staining was observed in all three cell lines treated with PI3K pathway inhibitors. Staining with clone D9E was a little less sensitive than with clone 14-5, but significant reduction or ablation of staining in cell lines treated with PI3K inhibitors was also observed. In summary, clones 14-5 and D9E appear to demonstrate adequate specificity and sensitivity for use in IHC assays.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 304.
Abstract 3042: Investigating microRNA expression profiles in colon cancer cells in response to cyclooxygenase 2 signaling
Authors: Harpster Kelsey L. , Bean Ronald R. , Jakobi Rolf , Chang Yu-Wen E.
Published: February 2011

MicroRNAs (miRNAs) are emerging as central players in cancer progression. Aberrant expression of miRNAs leads to malignant transformation and correlates with staging, progression and prognosis of cancers. Unfortunately, signaling pathways leading to altered expression of miRNAs are largely unknown. During early development of colon cancer, the expression of cyclooxygenase 2 (COX-2) is constitutively induced. We and others have shown that constitutive expression of COX-2 promotes colon carcinogenesis. However, the underlying mechanisms are not completely understood. To determine if COX-2 signaling alters expression of miRNAs, we used the Human Cancer RT2-miRNA PCR arrays to profile miRNA expression in HCA-7 human colon cancer cells treated with or without the COX-2 selective inhibitor NS-398. HCA-7 cells constitutively express endogenous COX-2. We found that expression levels of 39 miRNAs are different in HCA-7 cells as compared to HCA-7 cells treated with NS-398; 8 miRNAs were up-regulated and 31 miRNAs were down-regulated. To confirm the data from the PCR array, we used quantitative RT-PCR to quantify expression of 8 miRNAs in HCA-7 cells with or without NS-398 treatment. These 8 miRNAs were selected due to their relevance to cancer invasion and metastasis. We also compared the expression of these miRNAs in HCT-15 human colon cancer cells that were engineered to stably express COX-2 (HCT-15-COX-2) to the parental HCT-15 cells. Expression levels of 6 of the 8 examined miRNAs (miR-23b, 124, 146b-5p, 155, 183, 184) were significantly lower in HCT-15-COX-2 or HCA-7 cells than in parental HCT-15 or HCA-7 cells that were treated with NS-398. Therefore, we identified 6 miRNAs that are modulated by COX-2 and may play a role in mediating COX-2 signaling to promote colon carcinogenesis.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3042.
Abstract 3047: 2-Methoxyestradiol inhibits proliferation and induces apoptosis through modulation of miR-7/FLIP signaling in prostate cancer cells
Authors: Kumar Saikartik , Ghosh Rita , Kumar Addanki P.
Published: February 2011

Prostate cancer is the second most common cause of death related to cancer in Western societies. 2-Methoxyestradiol (2-ME2), an endogenous metabolite of 17-β estradiol inhibits tumor cell proliferation in various cancer cells, including in the prostate. Previous studies from our laboratory showed that 2-ME2 (i) inhibits proliferation of both androgen responsive and independent cells through induction of apoptosis involving G2/M check point block; (ii) prevents the development of prostate tumors in transgenic adenocarcinoma of mouse prostate (TRAMP) model. Although various molecular targets have been proposed, the mechanism of action behind its antiproliferative activity is still uncertain. Here we investigated the hypothesis that 2-ME2 may induce apoptosis in prostate cancer cells through modulation of microRNA (miRNAs). MicroRNAs are small non-coding RNAs that down regulate gene expression during cellular processes including proliferation and apoptosis. These studies led to the identification of miR-7 and −10 as predominantly up-regulated in response to 2-ME2 treatment in prostate cancer cell lines. Knock-down of miR-7 and −10 using mercury LNA probes showed resistance to 2-ME2-induced proliferation of prostate cancer cells. Additional studies to assess the effect of miR-7 knock-down to regulate apoptosis gene expression identified FLIP (FLICE -inhibitory protein) as one of the downstream targets of miR-7. We have shown that high grade prostate tumors express FLIP at higher levels compared to low grade and inhibition of prostate tumor development by 2-ME2 in preclinical animal model is associated with down regulation of FLIP. In addition overexpression of FLIP has been shown to contribute to resistance to apoptosis. We interpret our findings to suggest that up regulation of miR-7 following treatment with 2-ME2 inhibits FLIP leading to reduced proliferation and induction of apoptosis. Therefore our results may provide a molecular basis for the ability of 2-ME2 to suppress proliferation and to induce apoptosis in prostate cancer cells through modulation of miR-7 mediated inhibition of antiapoptotic genes including FLIP. Supported by ACS RSG-04-169 and NIH CA 135451 (APK).

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3047.