bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1365: Sun protection practices among melanoma survivors.
Authors: Puthumana Jeremy , Ferrucci Leah , Mayne Susan , Lannin Donald , Chagpar Anees
Published: August 2014

Purpose: Unprotected sun exposure is a key risk factor for melanoma and further exposure to ultraviolet radiation after a melanoma diagnosis may increase the risk of a subsequent melanoma. We sought to determine whether melanoma survivors observed sun protection practices more vigilantly than the rest of the population.

Methods: The National Health Interview Survey (NHIS) is an annual, cross-sectional survey of the civilian, non-institutionalized population of the United States on a wide range of health topics. We used the 2010 NHIS data on self-reported history of melanoma, sun protection practices, and indoor tanning to determine if individuals with a previous history of melanoma adhered more to sun protection practices than the general population.

Results: In 2010, the NHIS surveyed 27,120 adults, of whom 171 (0.74%) had a prior history of melanoma. 54.8% of melanoma survivors were men, and 10.2% were under the age of 40. More melanoma survivors reported they always stay in the shade than the general population (15.6% vs. 10.5%, p<0.001); however, 15.4% of melanoma survivors still reported rarely or never staying in the shade. Melanoma survivors were more likely (than their general population counterparts) to always wear a baseball cap/visor (31.3% vs. 18.4%, p=0.028), a wide-brimmed hat (20.5% vs. 6.1%, p<0.001), and a long-sleeved shirt (12.0% vs. 5.2%, p=0.003) when going outside on a warm, sunny day for more than one hour. They were also more likely to report always using sunscreen (32.0% vs. 17.2%, p=0.005); however, 27.3% of melanoma survivors reported never wearing sunscreen when going outside on a warm sunny day for more than one hour. No significant relationship was found between melanoma survivors and the general population in wearing long pants/skirts on a sunny day (22.2% of melanoma survivors always wear such clothing versus 17.7% of the general population, p=0.404). In addition, melanoma survivors were less likely to have used indoor tanning devices in the past 12 months than the general public (2.1% vs. 5.5%, p=0.009).

Conclusion: Melanoma survivors, in general, engage in more sun protection practices than the general population. However, the fact that many do not seek shade and never use sunscreen, and some reported tanning bed use, remains a concern for their skin cancer risk.

Citation Format: Jeremy Puthumana, Leah Ferrucci, Susan Mayne, Donald Lannin, Anees Chagpar. Sun protection practices among melanoma survivors. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1365. doi:10.1158/1538-7445.AM2013-1365
Abstract 1393: Fine mapping and characterization of Emca8.2, a genetic determinant of susceptibility to estrogen-induced mammary cancer on chromosome 5 in the rat.
Authors: Colletti John A. , Seiler Nicole L. , Hickman Maureen P. , Samanas Nyssa B. , Warren Chrsitopher L. , Shull James D.
Published: August 2014

Although estrogens have long been implicated in the etiology of breast cancer, the mechanisms through which estrogens contribute to breast cancer development remain poorly defined. Furthermore, many of the genes that influence the risk of developing breast cancer also remain to be determined. The ACI rat model of 17β-estradiol (E2)-induced mammary cancer is being used to elucidate the mechanisms through which estrogens contribute to breast cancer development and to identify novel genetic determinants of breast cancer susceptibility. Female ACI rats develop mammary cancer at an incidence nearing 100% when treated with physiological levels of E2. In contrast, the Brown Norway (BN) rat strain is resistant to E2-induced mammary cancer. A total of seven quantitative trait loci (QTL), designated Emca3 (Estrogen-induced mammary cancer) through Emca9, have been mapped in reciprocal intercrosses between ACI and BN rats. The purpose of the current study is to fine map the Emca8 genetic determinants of mammary cancer susceptibility, which was mapped to rat chromosome 5 (RNO5). ACI alleles at this QTL act to increase the number of mammary cancers per rat and to decrease the latency to the appearance of the first palpable mammary cancer. A series of congenic rat strains have been generated by introgressing defined segments of RNO5 from the resistant BN rat strain onto the genetic background of the susceptible ACI strain. By defining the susceptibility of these congenic rat strains relative to that of the parental ACI strain, a locus determining susceptibility to E2-induced mammary cancer, designated Emca8.2, has now been localized to a 1.1 megabase (Mb) region of distal RNO5 containing approximately 1200 genetic sequence variants. To help identify the mechanisms through which Emca8.2 determines susceptibility to mammary cancer, the expression levels of the candidate genes within the fine-mapped region have been investigated. The mapping and expression data that define the genomic architecture of Emca8.2 will be presented and discussed.

Citation Format: John A. Colletti, Nicole L. Seiler, Maureen P. Hickman, Nyssa B. Samanas, Chrsitopher L. Warren, James D. Shull. Fine mapping and characterization of Emca8.2, a genetic determinant of susceptibility to estrogen-induced mammary cancer on chromosome 5 in the rat. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1393. doi:10.1158/1538-7445.AM2013-1393
Abstract 1401: Overexpression of HER4 isoforms in transgenic mice reveal isoform-specific role in mammary gland development and carcinogenesis.
Authors: Wali Vikram B. , Gilmore-Hebert Maureen , Elenius Klaus , Stern David F.
Published: August 2014

Human epidermal growth factor receptor 4 (HER4/ERBB4) belongs to the family of four receptor tyrosine kinases (HER1-4). While HER1, HER2 and HER3 are often overexpressed in breast cancer and predict poor prognosis, the role of HER4 in cancer is still unclear. Some studies suggest a tumor suppressor role of HER4, while contradictory reports suggest its oncogenic potential. HER4 gene produces multiple isoforms as a result of alternative splicing, in contrast to other members of this receptor family. We have used a gain-of-function MMTV-HER4 model isoforms, to evaluate potential developmental and carcinogenic roles of HER4-CYT1 and CYT2. Mammary gland ductal morphogenesis was significantly suppressed by expressing one of the transgene isoforms, strongly suggesting a role of HER4 in normal mammary ductal morphogenesis in addition to its role in lactation described previously. Interestingly, sustained expression of one HER4 transgene isoform resulted in formation of mammary tumor lesions, whereas no significant findings were observed in the control group. Formation of tumor lesions in these transgenic mice strongly suggests an oncogenic function of a HER4 isoform. These novel findings may help improve the rational design of targeted breast cancer therapy. Supported by NIH grant RO1 CA80065.

Citation Format: Vikram B. Wali, Maureen Gilmore-Hebert, Klaus Elenius, David F. Stern. Overexpression of HER4 isoforms in transgenic mice reveal isoform-specific role in mammary gland development and carcinogenesis. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1401. doi:10.1158/1538-7445.AM2013-1401
Abstract 143: Breast cancer incidence rate in american women started to increase: Trend analysis from 2000 to 2009.
Authors: Hou Ningqi , Huo Dezheng
Published: August 2014

Background: Recent report showed that breast cancer incidence rate in the US stabilized after a sharp reduction in 2002 and 2003. It is important to continue monitoring breast cancer incidence rates according to age group, race/ethnicity, estrogen receptor (ER), and tumor stage.

Methods: Age-standardized breast cancer incidence rates were calculated using data from the Surveillance, Epidemiology, and End Results (SEER) 18 registries from 2000 to 2009, among 677,774 female breast cancer patients aged 20 and above. Jointpoint regression models were used to fit a series of joined straight lines on a log scale to the trends of annual age-standardized rates.

Results: The overall breast cancer incidence significantly increased for non-Hispanic black from 2005 to 2009 (annual percentage change, APC =2.0%, p=0.01) and Asian/Pacific Islanders from 2000 to 2009 (APC 1.2%, p=0.02). Since 2004, incidence rates in women age 40-49 significantly increased for all racial/ethnic groups (overall APC = 1.1%, p=0.001). Incidence rate of carcinoma in situ significantly increased in all racial/ethnic groups (APC ranges from 2.3%-3.0%, p<0.005) and localized breast cancer incidence significantly increased in non-Hispanic blacks (APC=1.3%, p=0.004) and Asian (APC=1.2%, p=0.03). ER positive breast cancer significantly increased in almost all age/race sub-groups after 2005 (APC by race: non-Hispanic white 1.5%, non-Hispanic black 4.3%, Asian/Pacific Islander 1.7%, and Hispanic 1.8%, p-values<0.05), while ER negative breast cancer decreased in most sub-groups (APC by race: non-Hispanic white -3.9%, non-Hispanic black -3.7%, Asian/Pacific Islander -1.5%, and Hispanic -4.3%, p-values<0.05).

Conclusions: Recent incidence of breast cancer appears to be increasing in certain subgroups, including non-Hispanic black, Asian/Pacific Islanders, women 40-49 years old, low-stage breast cancer, and estrogen-receptor positive breast cancer. These changes are unlikely due to mammography screening rate that has remained stable in recent years, but may be partially explained by the transition from film to digital mammography, which has higher sensitivity to detect breast cancer, especially young-onset breast cancer. It deserves further investigation on why the rate of estrogen receptor negative breast cancer declined.

Citation Format: Ningqi Hou, Dezheng Huo. Breast cancer incidence rate in american women started to increase: Trend analysis from 2000 to 2009. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 143. doi:10.1158/1538-7445.AM2013-143
Abstract 1463: Detection of cancer before distant metastasis.
Authors: Coumans Frank AW , Siesling Sabine , Terstappen Leon WMM
Published: August 2014

Introduction: The majority of deaths from cancer are due to disseminated disease rather than the primary tumor. As metastases are often discovered years after surgical removal of the primary tumor, probably at least one metastasis was already present at the time of surgery. To establish a distant metastasis cells must disseminate from the primary tumor and overcome a series of obstacles, the metastatic cascade. In this study we develop a mathematical model for this cascade to estimate the technology requirements for detection of a primary tumor before the first metastasis has been formed.

Methods: The metastatic cascade is described in discrete steps: 1. local tumor growth; 2. dissemination into circulation; 3. survival in circulation; 4. extravasation into tissue; and 5. growth into a metastasis. The model was built using data and relationships described in the literature to predict the relationship between tumor size and probability of distant metastasis for 38715 patients with surgically removed TxNxM0 primary breast cancer from the Netherlands Cancer Registry. The model was calibrated using primary tumor size, probability of distant metastasis and time to distant metastasis for 1489 patients with stage T1bNxM0 (25% of total patients with T1bNxM0). Validation of the model was done with data for all patients.

Results: From the time to distant metastasis of these 38715 breast cancer patients, we determined a tumor doubling time of 1.7 ± 0.9 months. Fitting the data for 25% of T1b patients estimates a metastatic efficiency of 1 metastasis formed per 60 million disseminated tumor cells. Validation of the model to data of patients in all T-stages shows good agreement between model and epidemiological data. To reduce the 5-year risk of distant metastasis for TxNxM0 from 9.2% to 1.0%, the primary tumor needs to be detected and removed before it reaches a diameter of 2.7 ± 1.6 mm. At this size, the model predicts that there will be 9 ± 6 CTC/L blood.

Conclusions: To reduce the rate of distant metastasis in surgically treated TxNxM0 breast cancer to 1%, imaging technology will need to be able to detect lesions of 2.7 mm in diameter or smaller. Before CTC detection can be applied in the early disease setting, sensitivity will need to be improved by at least 15-fold and combined with technology that minimizes false positives.

Citation Format: Frank AW Coumans, Sabine Siesling, Leon WMM Terstappen. Detection of cancer before distant metastasis. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1463. doi:10.1158/1538-7445.AM2013-1463
Abstract 1464: In vitro culturing of CTCs/DTCs in personalized cancer management.
Authors: Bobek Vladimir
Published: August 2014

Subsequent detection, quantification and molecular characterization of CTCs has an enormous potential in numerous of oncology areas. But detection and isolation of CTCs is technically quite challenging. The whole dissemination process encompasses also individual tumor cells which are found in relatively big quantities in peritoneal washings (PW) and pleural effusions (PE), so called disseminated tumor cells (DTCs).

Thanks in vitro cultures of separated CTCs/DTCs we were able to introduce additional chemosensitivity testing for the purpose of individualized cancer treatment. We present an isolation and cultivation of CTC/DTC of non-small cell lung cancer and gastrointestinal cancers (esophageal cancer, colorectal cancer, gastric cancer, pancreatic cancer) describing cultivation methodology on patients case studies.

The cancer cells were separated from body fluids (PW, PE, blood) using the differences in size of cancer cells if compared to other nucleated cells detected in body cavities. After reaching the confluence the chemosensitivity of cell cultures has been tested by xCELLigence system (RTCA, Roche) for different concentrations of single cis-platinum, gemcitabine and fluorouracyl (incubation time 24-72 hours) or their combinations. The dynamic real-time cell growth monitoring enables parallel or sequential combinations of administered drugs with a direct outcome for patients care.

Conclusions: We assume that developing cultivation strategies for CTCs/DTCs could bring us closer to the definition of the CTC/DTC-function within the dissemination process with a perspective to use this knowledge in personalized cancer treatment.

Citation Format: Vladimir Bobek. In vitro culturing of CTCs/DTCs in personalized cancer management. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1464. doi:10.1158/1538-7445.AM2013-1464
Abstract 1480: The effect of Metformin on microtentacle formation and stabilization in breast cancer cells.
Authors: Chakrabarti Kristi , Whipple Rebecca A. , Boggs Amanda E. , Charpentier Monica S. , Slovic Jana , Vitolo Michele I. , Martin Stuart S.
Published: August 2014

Metastasis is one of the main causes of mortality in women with breast cancer. Epithelial-to-mesenchymal transition (EMT) has emerged to be a key mediator of breast cancer metastasis. It has previously been shown that EMT can stabilize and increase microtentacles (McTNs) in breast cancer cell lines and enhance their metastatic potential. McTNs are dynamic, microtubule-enriched plasma membrane extensions that are stabilized with the exposure of a carboxy-terminal glutamic acid residue in alpha-tubulin (Glu-tubulin). McTN stabilization has been shown to promote tumor cell aggregation and reattachment in vitro. In this study, we investigated the effect of EMT suppression on McTN formation and stabilization in breast cancer cells by the dimethyl biguanide, Metformin. While it is widely used to treat type 2 diabetes, Metformin has more recently been shown to inhibit EMT in multiple breast cancer cell lines. We show here that Metformin decreased mesenchymal markers, such as N-cadherin and vimentin, in a dose-dependent manner in MDA-MB-231 breast cancer cells. Confocal and fluorescent microscopy was used to visualize and quantify McTN formation in these cells. There was a significant decrease in McTNs in MDA-MB-231 cells treated with 5mM Metformin for 24 hours. We correlated the decrease in the number of McTNs with a decrease in Glu-tubulin protein levels, which suggests that Metformin also inhibits McTN stabilization. The decrease in McTN formation and stabilization was also associated with diminished tumor cell reattachment in vitro. In addition, Metformin was able to protect MDA-MB-231 cells from increased McTN formation induced by the microtubule stabilizing agent, Taxol. Metformin's mechanism of action on breast cancer cells to elicit these cytoskeletal modifications has yet to be elucidated. However, these results suggest a role for Metformin at multiple stages of tumor progression to protect breast cancer patients from clinical spread of disease.

Citation Format: Kristi Chakrabarti, Rebecca A. Whipple, Amanda E. Boggs, Monica S. Charpentier, Jana Slovic, Michele I. Vitolo, Stuart S. Martin. The effect of Metformin on microtentacle formation and stabilization in breast cancer cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1480. doi:10.1158/1538-7445.AM2013-1480
Abstract 1488: Metastatic analysis of clonal epithelial and mesenchymal-like murine mammary tumor cells.
Authors: Palen Katie A. , Johnson Bryon D. , Gershan Jill A.
Published: August 2014

Tumor metastasis is the most common cause of cancer mortality, and yet it is among the most poorly understood processes in tumor progression. While carcinomas are composed primarily of epithelial cells, tumors contain a continuum of cells with epithelial and mesenchymal-like properties. This tumor cell heterogeneity is attributed to differentiation plasticity, which is characterized by epithelial to mesenchymal (EMT) or mesenchymal to epithelial transitions (MET). It has been proposed that tumor cells undergo EMT in order to disseminate from the primary tumor and MET in order to seed at secondary sites. The invasive mesenchymal carcinoma cell phenotype is thought to be a state required for tumor dissemination, but to date, pathologists have not found conclusive evidence of mesenchymal cells in human metastatic tumor samples. An alternative theory suggests that both mesenchymal and epithelial cells are required for metastasis to occur. According to this theory, detachment from the primary tumor and invasion by mesenchymal cells facilitates the movement of epithelial tumor cells that are able to seed at distant sites. We hypothesize that the presence of mesenchymal-like mammary tumor cells facilitate the metastasis of epithelial tumor cells and that only epithelial cells are able to seed at distant sites. To test this hypothesis, we cloned epithelial and mesenchymal-like primary cell lines from spontaneous mammary tumors produced in FVB MMTV/Neu mice. Epithelial cells have been stably transfected to express firefly luciferase and GFP, and we are in the process of cloning mesenchymal-like cells expressing Renilla luciferase and mCherry. We have preliminary data that shows that when clonal epithelial cells are administered into the mammary fat pad of FVB MMTV/Neu mice, tumors grow, but do not form metastatic lesions. In contrast, when stably transfected clonal epithelial cells are administered with non-transfected clonal mesenchymal-like tumor cells, luciferase+ metastatic lesions in the spleen, liver and lung are detected biophotonically, and GFP + cells are detected in the spleen and liver. These data indicate that epithelial cells are present (but does not rule out the presence of mesenchymal cells) at the metastatic site. Whether epithelial cells undergo EMT, and how metastasis is facilitated by the presence of mesenchymal cells remains to be determined.

Citation Format: Katie A. Palen, Bryon D. Johnson, Jill A. Gershan. Metastatic analysis of clonal epithelial and mesenchymal-like murine mammary tumor cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1488. doi:10.1158/1538-7445.AM2013-1488
Abstract 1491: Downregulation of KLF5 contributes to TGF-β- and EGF-induced EMT by reducing miR-200s.
Authors: Zhang Baotong , Dong Jin-tang
Published: August 2014

Krüpple-like factor 5 (KLF5) is a basic transcriptional factor that has been well documented for its regulatory role in the proliferation and survival of epithelial cells. Recently, bioinformatic analysis suggests that KLF5 also regulates epithelial-to-mesenchymal transition (EMT), yet experimental evidence is still lacking regarding whether and how KLF5 regulates EMT. As a form of epithelial cell plasticity, EMT can be induced by many cytokines including TGF-β and EGF. Here we report a role of KLF5 in EMT. We found that KLF5 was significantly down-regulated during TGF-β- and EGF-induced EMT in epithelial cells including HaCaT cells, MCF-10A cells, and mouse primary keratinocytes; and that silencing KLF5 induced EMT as well, as indicated by altered cell morphology, enhanced migration, upregulation of CDH2, FN1 and ZEB1, dislocation of CDH1 from cell-cell junctions, and induction of actin fibers. To determine whether miRNAs mediate the role of KLF5 in EMT, a real-time qPCR assay was employed to search for miRNAs regulated by KLF5 during EMT. Five members of the miR-200 family, all of which have been implicated in EMT, were found to be down-regulated upon the downregulation of KLF5. KLF5 directly regulated the expression of miR-200s by binding to the GC boxes of their promoters, as demonstrated by functional and biochemical analyses including luciferase reporter assay, ChIP-PCR assay, and oligo pull-down assay. Functionally, overexpression of miR-200s inhibited EMT induced by KLF5 silencing or TGF-β and EGF treatment in HaCaT cells, and forced expression of KLF5 attenuated TGF-β- and EGF-induced EMT by rescuing miR-200s expression. To evaluate whether the KLF5-miR-200s axis modulates EMT in vivo, we determined the expression of Klf5, miR-200s and EMT markers in knockout mice and human breast cancer specimens. In mouse prostate neoplasms induced by Pten deletion, knockout of Klf5 repressed the expression of miR-200s and induced the downregulation of CDH1 and upregualtion of vimentin, which are indicative of EMT. Expression of KLF5 also correlated with miR-200s at the RNA level in human breast cancer specimens. These findings suggest that KLF5 directly regulates the transcription of miR-200s in epithelial cells, and that downregulation of the KLF5-miR-200 axis plays a causal role in TGF-β- and EGF-induced EMT, providing new insights into the regulation of EMT.

Citation Format: Baotong Zhang, Jin-tang Dong. Downregulation of KLF5 contributes to TGF-β- and EGF-induced EMT by reducing miR-200s. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1491. doi:10.1158/1538-7445.AM2013-1491
Abstract 1497: Inducing a mesenchymal-to-epithelial transition for the targeting of cancer stem cells.
Authors: Pattabiraman Diwakar R. , Reinhardt Ferenc , Bierie Brian , Weinberg Robert A.
Published: August 2014

The occurrence of metastasis and the persistence of chemotherapy-resistant carcinoma cells are the two leading factors that contribute to cancer-related deaths. Both these properties of tumors have been shown to arise from the epithelial-to-mesenchymal transition (EMT), a process that leads to the generation of cancer stem cells (CSCs). Hence, it is believed that eradication of these CSCs represents the most effective way to curb tumor progression and relapse. Recent studies from the Weinberg lab have implicated key signalling pathways in the induction and maintenance of the mesenchymal state, including TGFβ and Wnt. Through this project, we propose to use a number of small-molecule inhibitors of specific signaling pathways, including those mentioned above, to induce a mesenchymal-to-epithelial transition (MET) in human breast cancer cell lines. Upon inhibition of these essential pathways, the ability of these cells to form tumors, seed metastases, and exhibit resistant to doxorubicin and paclitaxel will be tested. Preliminary results show selective targeting of mesenchymal cells upon treatment with a combination of TGFβ and non-canonical Wnt inhibitors, which appear to induce their apoptosis as well as promote their differentiation back to an epithelial-like state. We hypothesize that upon inducing differentiation of the mesenchymal/stem-like population of the cancer cells using various combinations of small-molecule inhibitors, they will lose their ability to metastasize and will exhibit an increased susceptibility to apoptosis upon treatment with the above-mentioned chemotherapeutic agents. If successful, these inhibitors, or improved versions, will have the potential to be used in conjunction with chemotherapeutic drugs in the clinic to curb the relapse and progression of cancers. The significance of this work extends beyond breast cancers to a large number of carcinomas that have been shown to undergo an EMT, including prostate, colon and lung cancers.

Citation Format: Diwakar R. Pattabiraman, Ferenc Reinhardt, Brian Bierie, Robert A. Weinberg. Inducing a mesenchymal-to-epithelial transition for the targeting of cancer stem cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1497. doi:10.1158/1538-7445.AM2013-1497

Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.
Abstract 1545: Education of macrophages through modulation of NF-kappaB: an opportunity for targeted therapy.
Authors: Barham Whitney , Tikhomirov Oleg , Chen Lianyi , Ortega Ryan , Gleaves Linda , Onishko Halina , Sherrill Taylor , Connelly Linda , Blackwell Timothy S. , Yull Fiona E.
Published: August 2014

Macrophages are a plastic cell type, capable of adapting to numerous signals within their environment. As part of the innate immune system, macrophages were traditionally considered anti-tumor (M1), but it has been well established that macrophages can also help to create a pro-tumor, pro-metastatic tumor niche (M2). NF-κB transcription factors can regulate both pro- (MMP's, VEGF) and anti-tumor (iNOS) downstream targets within macrophages, suggesting that modulation of NF-κB may play a role in the two different macrophage phenotypes. However, our understanding of NF-κB signaling specifically within macrophages during tumor progression is limited. To this end, we have developed murine transgenic models that enable us to induce expression of an activator or dominant inhibitor of NF-κB in macrophages by adding doxycycline to the drinking water of mice. We have combined these novel transgenics with the polyoma model of mammary cancer for our studies.

We have recently shown that activation of NF-κB in macrophages significantly limits metastasis in a tail vein model of tumor progression. In this model, constitutive IKK2 activity within macrophages leads to an anti-tumor immune response including altered immune cell populations within the lung microenvironment, changes in chemokine and cytokine expression and rapid killing of tumor cells during the seeding phase mediated by reactive oxygen species. Our current work has extended these findings to an orthotopic mammary tumor model. Again, we find that activation of NF-κB in macrophages results in decreased primary tumor growth and decreased tumor seeding into the blood. To model this activation in vitro, we have utilized immortalized bone marrow derived macrophages from IκBα knock-out mice compared to a wild type line. IκBα KO macrophages display changes in morphology and adhesion relative to wild type macrophages. This correlates with increased cytotoxic behavior in co-culture with polyoma tumor cells, mirroring the in vivo phenotype of the cIKK2 expressing macrophages. Given these findings, we believe that targeted activation of NF-κB signaling in macrophages could be harnessed to overcome the education of macrophages by tumor cells, and could be exploited as a novel targeted therapy.

Citation Format: Whitney Barham, Oleg Tikhomirov, Lianyi Chen, Ryan Ortega, Linda Gleaves, Halina Onishko, Taylor Sherrill, Linda Connelly, Timothy S. Blackwell, Fiona E. Yull. Education of macrophages through modulation of NF-kappaB: an opportunity for targeted therapy. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1545. doi:10.1158/1538-7445.AM2013-1545
Abstract 1568: Evaluate the efficacy of new therapeutics using advanced metastatic mouse models.
Authors: Zhang Juan , Guo Sheng , Cai Jie , Sun Yanmei , Mao Guanping , Wang Zhun , Zhang Lan , Jin Ying , Shi Qian , Chen Taiping
Published: August 2014

Metastasis, the spread and growth of tumor cells to distant organs, is usually a final and fatal step in the progression of solid malignancies. Preclinical test of the new anti-cancer therapies that could mimic the disease present in most clinical trials asks for robust metastatic models than just the subcutaneous models. We sought to establish and validate a panel of experimental (i.v. inoculation) and spontaneous (orthotopic inoculation) mouse models with high metastatic potential and varied tissue tropism in different types of cancer models, covering colorectal, liver, prostate, breast, pancreatic cancer and melanoma. The responsiveness of metastasis to several therapies that are proved to be clinically effective or ineffective was investigated. A higher predictive power of the metastatic models was demonstrated compared with the corresponding subcutaneous models. By means of the transcriptomic analysis of the paired primary tumor population and the distant lung metastasis population in 5 orthotopic tumor models, we identified the gene signatures that differentiate the two populations in different tumor types. Our signature was also compared with the clinical relevant gene signatures correlated with lung metastasis risk. Several genes are concordantly expressed in both signatures and are biologically meaningful. Overall, given the higher predictive power and the clinical relevance, the use of the metastatic mouse models constitutes an effective and important strategy to preclinically evaluate the efficacy of new therapeutic regimens.

Citation Format: Juan Zhang, Sheng Guo, Jie Cai, Yanmei Sun, Guanping Mao, Zhun Wang, Lan Zhang, Ying Jin, Qian Shi, Taiping Chen. Evaluate the efficacy of new therapeutics using advanced metastatic mouse models. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1568. doi:10.1158/1538-7445.AM2013-1568
Abstract 1584: Targeting the mitotic checkpoint with inhibition of MPS1 kinase enhances radiosensitivity of glioblastoma cancer cells.
Authors: Tandle Anita T. , Hanson Ryan , Maachani Uday B. , Meushaw Tamalee , Zhao Shuping , Shankavaram Uma , Tofilon Philip , Caplen Natasha J. , Camphausen Kevin
Published: August 2014

During the cell cycle, genomic stability requires accurate chromosome segregation. Errors in this process can cause aneuploidy and lead to tumorigenesis. To ensure faithful chromosome segregation, cells develop a mechanism called the spindle assembly checkpoint (SAC). Cancer cells are addicted to the components of SAC machinery for a faithful entry of the cell into anaphase. Thus, targeting the molecular mechanisms required for the growth of aneuploid cells may be a more cancer cell specific therapeutic approach applicable to broader tumor histologies. Previously, using a siRNA based RNAi screen we identified MPS1 kinase, (also known as TTK) as an important kinase for GBM cell survival. MPS1 is an essential SAC enzyme aberrantly overexpressed in a wide range of tumors and necessary for tumor cell proliferation.

We observed inhibition of GBM cell growth when MPS1 was downregulated by number of MPS1 specific siRNAs. This was further validated using a selective and orally bioavailable MPS1 inhibitor NMS-P715 in various in-vitro cell assays. The inhibition of cell death was induced partly by apoptosis; however, the major mechanism was mitotic catastrophe. Cells treated with NMS-P715 showed an increase in cells in G2-M phase of cell cycle compared to control cells followed by mitotic catastrophe. Moreover, inhibition of MPS1 resulted in radiosensitization of GBM cells. We observed decrease in DNA damage repair and significant retention of γH2AX foci after combination of radiation (RT) with NMS-P715 compared to individual treatments. Next, radiation in combination with NMS-P715 inhibited cell survival ability of GBM cells in a colony formation assay. Further, NMS-P715 could inhibit GBM tumor growth in an orthotopic brain tumor model. Finally, in order to determine MPS1 associated molecular pathways, we compared gene expression profile in MPS1 knockdown cells compared to the control by microarray analysis. Ingenuity pathway and Gene Set Enrichment Analysis were used to investigate the biological relevance of the MPS1 modulated genes. We identified genes important in cell assembly, cell organization, DNA repair and cell death pathways. Thus, inhibiting MPS1 kinase in combination with radiation could represent a promising new approach to GBM therapy.

Citation Format: Anita T. Tandle, Ryan Hanson, Uday B. Maachani, Tamalee Meushaw, Shuping Zhao, Uma Shankavaram, Philip Tofilon, Natasha J. Caplen, Kevin Camphausen. Targeting the mitotic checkpoint with inhibition of MPS1 kinase enhances radiosensitivity of glioblastoma cancer cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1584. doi:10.1158/1538-7445.AM2013-1584
Abstract 1614: Murine matrigel plug assay for quick evaluation of anti-angiogenesis drugs.
Authors: Sun Yanmei , Song Xiaoming , Li Nan , Cai Jie , Shi Qian , Chen Taiping
Published: August 2014

Angiogenesis plays important roles in disease processes such as tumor development and progression, as well as many physiological conditions. A few cancer drugs were successfully approved based on anti-angiogenesis mechanisms. More candidates are in different evaluation stages, including clinical phase II stages. Among many assays used for angiogenesis evaluation, Murine Matrigel Plug Assay (MMPA) is a relatively simple, highly effective in vivo assay to evaluate anti-angiogenesis responses of the test articles. Because of its simplicity and short assay duration, the assay is scalable for efficient screening of large number of test articles. However, the analysis of the plugs, usually with H&E staining and microvessel counting, is time consuming and error prone, hampers the usage of this method as a general screening assay for test articles. Here, we set out to test a number of combinations of assay conditions, including mouse strains, angiogenesis inducers, varying days of treatment, and different routes of SOC drug treatments. More importantly, we tested plug analysis methods including H&E staining, CD31 IHC staining, as well as hemoglobin concentration analysis. We demonstrated that hemoglobin concentration analysis results are corroborated by IHC staining as well as H&E staining, in most of the conditions tested. Hemoglobin assay is less time consuming and easier to scale up. Thus MMPA followed by hemoglobin analysis may be suitable for large scale screening studies for angiogenesis modulators. Interestingly, CTX, one of the popular anti-cancer therapeutics, was shown in MMPA as a good anti-angiogenesis agent. In conclusion, the MMPA described may greatly facilitate the preclinical development of cancer therapeutics that target the angiogenesis.

Citation Format: Yanmei Sun, Xiaoming Song, Nan Li, Jie Cai, Qian Shi, Taiping Chen. Murine matrigel plug assay for quick evaluation of anti-angiogenesis drugs. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1614. doi:10.1158/1538-7445.AM2013-1614
Abstract 1619: Combination of VEGFR inhibitor lenvatinib (E7080) and Met/EphB4 inhibitor golvatinib (E7050) overcomes VEGFR inhibitor-resistant tumor vascular.
Authors: Kawano Satoshi , Nakagawa Takayuki , Nakazawa Yuya , Sato Yoshiaki , Matsushima Tomohiro , Funahashi Yasuhiro
Published: August 2014

Vascular endothelial growth factor receptor (VEGFR) inhibitors are approved for the treatment of several types of tumors, however, some tumor shows intrinsic resistance to VEGFR inhibitors. In addition, patients initially responding develop acquired resistance to VEGFR inhibitors after varying periods of time. Thus it is urgent to develop strategy to overcome anti-VEGF therapy resistance. Hepatocyte growth factor (HGF)/Met signaling is activated in various types of tumors. Recently, it is reported that HGF/Met signaling contributes to resistance to tyrosine kinase inhibitors such as EGFR, BRAF, ALK inhibitors. This suggests that Met inhibitors may be promising in treating HGF/Met activated anti-VEGF therapy-resistant tumors. We have identified lenvatinib (E7080) as a highly-potent VEGFR inhibitor and phase III trial is ongoing in thyroid cancer. Lenvatinib inhibited the growth of HUVEC induced by VEGF, however, addition of HGF to HUVEC conferred resistance to lenvatinib. This resistance was cancelled by the addition of golvatinib (E7050) which is identified as a potent Met/EphB4 inhibitor and phase I trial of the lenvatinib/golvatinib combination therapy is ongoing. In mouse models bearing various tumor cell lines expressing VEGF and HGF, treatment with lenvatinib alone demonstrated only moderate antitumor activity, however, combination with golvatinib demonstrated synergistic antitumor activity with decrease of tumor vessel density. Furthermore, it is reported that EphB4 has critical roles in tumor vessel development and maturation. Our unique in vitro assay systems, endothelial cells/pericyte 2D co-culture assay and endothelial cells/pericyte 3D co-culture sprouting assay, suggested that EphB4 inhibitory activity of golvatinib disrupted anti-VEGF therapy-resistant tumor vascular in the combination with lenvatinib. Together, these data demonstrated that combination therapy with golvatinib may be promising to overcome anti-VEGF therapy resistance.

Citation Format: Satoshi Kawano, Takayuki Nakagawa, Yuya Nakazawa, Yoshiaki Sato, Tomohiro Matsushima, Yasuhiro Funahashi. Combination of VEGFR inhibitor lenvatinib (E7080) and Met/EphB4 inhibitor golvatinib (E7050) overcomes VEGFR inhibitor-resistant tumor vascular. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1619. doi:10.1158/1538-7445.AM2013-1619
Abstract 1621: Short-time exposure to high concentrations of sunitinib causes tumor cell death.
Authors: Rovithi Maria , Poel Dennis , Voortman Jens , Labots Mariette , Dekker Henk , Gotink Kristy J. , Broxterman Henk J. , Verheul Henk M.W.
Published: August 2014

Background: Sunitinib, although initially developed as an antiangiogenic agent, has been shown to exert direct anti-tumor effects. Previously we demonstrated that sunitinib inhibits proliferation of various tumor cell lines in a dose-dependent manner. We here studied whether short exposure to high peak concentrations of sunitinib would induce tumor cell death.

Methods: 786-O renal cancer cells were incubated with increasing concentrations of sunitinib (5, 10 and 20 μM) for various time intervals (1 hour, 3, 6, 9, 24, 48, 72 hours). At prespecified time points, sunitinib was removed and cells were subsequently allowed to grow in drug-free culture medium. Viability of these cells one week after short time sunitinib exposure and sensitivity of recovered cells after repetitive treatment with high sunitinib concentrations were determined by MTT assays. Flow cytometry with Propidium Iodide staining was employed to determine apoptosis.

Results: Short-time exposure to high concentrations of sunitinib significantly induced cell death. Exposure to 20 μM of sunitinib for 9 hours provoked cell death in a manner identical to the prolonged exposure of 24, 48 or 72 hours. Flow cytometry confirmed the significant increase in cell death (60%) after 9-hour treatment with 20 μM sunitinib, compared to the untreated control (2%). Tumor cells that were repetitively treated with high concentrations of sunitinib retain their sensitivity to the drug identical to its parental cell line. Analysis of the key signaling pathways involved in sunitinib-induced cell death is ongoing.

Conclusion: We here report that repetitive short-time exposure to high concentration sunitinib causes cell death without inducing resistance. Interestingly, single-dose sunitinib up to 300 mg was previously reported to be well tolerated by patients. Based on these findings, administration of high dose intermittent sunitinib may result in cytotoxic antitumor activity irrespective of angiogenesis inhibition. A phase I trial to test this hypothesis in patients with cancer will start shortly.

Citation Format: Maria Rovithi, Dennis Poel, Jens Voortman, Mariette Labots, Henk Dekker, Kristy J. Gotink, Henk J. Broxterman, Henk M.W. Verheul. Short-time exposure to high concentrations of sunitinib causes tumor cell death. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1621. doi:10.1158/1538-7445.AM2013-1621
Abstract 1625: Crosstalk between breast cancer and stromal cells occurs via exosomes and gap junctions in bone metastatic breast cancer.
Authors: Bussard Karen M. , Marini Frank C.
Published: August 2014

Breast cancer preferentially metastasizes to the bone, where the five-year relative survival rate falls from 90% to <10%. Although the precise mechanism underlying preferential metastasis is unknown, the bone likely provides a hospitable environment that both attracts breast cancer cells and allows them to colonize and grow. Besides affects osteoblast and osteoclast properties, we have evidence that metastatic breast cancer cells further create a unique bone niche by orchestrating extensive crosstalk with osteoblasts that may be involved in creating a niche permissive for breast cancer cell dormancy.

MC3T3-E1 murine osteoblasts were grown to confluence, then MDA-MB-231 human breast cancer cells added at a ratio of 1 breast cancer cell to 10 osteoblasts. Co-cultures were subsequently fixed and stained for Cx43 expression. Next, MC3T3-E1 cells and MDA-MB-231 cells were grown to confluence. Culture supernatants were removed and processed through a series of ultra-centrifugation, purification, and concentration steps to purify for exosomes. Finally, MC3T3-E1 cells were grown to various stages of maturity: 4 (growth) and 10 days (early differentiation) and incubated with conditioned medium from human MDA-MB-231 cells. qPCR was used to assay for changes in osteoblast-derived inflammatory cytokine expression.

Expression of the gap junction protein Cx43 was localized between breast cancer cells and osteoblasts. Both MC3T3-E1 and MDA-MB-231 cells expressed exosomes, which were visualized using a transmission electron microscope. Osteoblast-derived cytokines increased in the presence of MDA-MB-231 breast cancer cell conditioned medium included MCP-1 (CCL2), LIX (CXCL5), LIF, and NFATc2.

Overall, these data suggest that suggest there is extensive crosstalk directly between osteoblasts and breast cancer cells, and that osteoblasts are an important source of cytokines in breast cancer bone metastasis. Thus, these findings implicate the bone microenvironment and cancer cell manipulation thereof in orchestrating metastatic tumor cell dormancy, colonization, and survival. Supported in part by the National Institutes of Health; 1RC1 CA146381, 1R01NS06994, P50 CA083639 for FCM; Ruth L. Kirschstein Institutional Research Service Award (NRSA) T32 CA079448 for KMB.

Citation Format: Karen M. Bussard, Frank C. Marini. Crosstalk between breast cancer and stromal cells occurs via exosomes and gap junctions in bone metastatic breast cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1625. doi:10.1158/1538-7445.AM2013-1625
Abstract 1635: MiR-21/Smad 7 signaling determines TGF-β1-induced cancer associated fibroblast (CAF) formation.
Authors: Li Qiong , Wang Yongbin , Xiong Wujun , Mi Jun
Published: August 2014

Tumor-secreted growth factors such as TGF-β1 transform adjacent normal fibroblasts into cancer-associated fibroblasts (CAF) which is a major source of CAF. TGF-β1 has canonical and noncanonical Smad pathway, which pathway mediates TGF-β1 signaling during the CAF formation and how TGF-β1 activates regulatory Smad proteins is not entirely understood. To investigate whether the miR-21/Smad 7 signaling determines TGF-β1-induced CAF formation, the in vitro TGF-β1-induced CAF model was successfully established and the function of fibroblasts knocked down or overexpressed miR-21 or Smad 7 were finally identified in the xenografted mice, with or without TGF-β1 stimulation. Our data showed that the mature miR-21 is increased in fibroblast after TGF-β1 treatment, and miR-21 regulates Smad 7 protein level through the translation inhibition instead of the mRNA decay. Moreover, our data showed Smad 3 was phosphorylated 2 hours early than Smad 2 after TGF-β1 stimulation and Smad

7 knockdown enhanced Smad 2 and 3 phosphorylation; both of their phosphorylations were abolished by the TGFR kinase inhibitor. Most important, both of the over-expression of miR-21 or the knockdown of Smad 7 promoted CAF formation, even without TGF-β1 stimulation while the miR-21 knockdown or the Smad 7 overexpression as well as Smad 2, 3 and Smad 4 knock down blocked the TGF-β1-induced CAF formation.

These observations suggest that TGF-β1 induces CAF formation through the canonical pathway, miR-21/Smad 7 is a critical mediator.

Citation Format: Qiong Li, Yongbin Wang, Wujun Xiong, Jun Mi. MiR-21/Smad 7 signaling determines TGF-β1-induced cancer associated fibroblast (CAF) formation. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1635. doi:10.1158/1538-7445.AM2013-1635

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