bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1488: SOX2 promotes tumor metastasis by stimulating the process of epithelial-mesenchymal transition
Authors: Li Xuefei , Jia Xianpei , Xu Yingxi , Chen Yanan , Chen Si , Xiang Rong , Li Na
Published: June 2012

Ectopic expression of SOX2 has been reported in a variety of tumor tissues and contributes to cell proliferation and tumorigenesis by transcriptional regulation of the target genes; however the role of SOX2 in tumor metastasis and the underlying mechanisms remain largely elusive. Here we used immunohistochemistry to analyze the expression of SOX2 in the tissues of human breast and lymph nodes and found that SOX2 was highly expressed in breast cancer tissues and the metastasised lymph node, also the expression of SOX2 was closely correlated with the TNM stage and histological grade. We further demonstrated that SOX2 improved the migration and invasion properties of breast cancer and prostate cancer cells by facilitating the process of epithelial-mesenchymal transition (EMT). Investigation of the molecular mechanism revealed that SOX2 regulated EMT by β-catenin signaling pathway. Our findings prove that SOX2 plays a pivotal role in the metastasis of human breast cancer and prostate cancer and suggest that SOX2 has the potential to be developed as the diagnosis marker for breast and prostate cancer metastasis.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1488. doi:10.1158/1538-7445.AM2011-1488
Abstract 1500: A novel nutrient mixture exhibits antitumor activity in human fibrosarcoma cell line HT-1080
Authors: Roomi M. Waheed , Jariwalla Neil , Roomi Nusrath , Rath Matthias , Niedzwiecki Aleksandra
Published: June 2012

Introduction: Fibrosarcoma, an aggressive and highly metastatic cancer of connective tissue, is generally associated with poor prognosis. Cancer mortality usually results from tumor invasion of local tissue and metastasis to vital organs. We investigated the effect of a specific nutrient mixture (PB) of quercetin, cruciferex, curcumin, green tea extract and resveratrol on human fibrosarcoma cell line HT-1080 for viability, MMP expression, invasion, apoptosis and morphology.

Material and Methods: Human fibrosarcoma cell line HT-1080 (ATCC) was cultured in DEM media, supplemented with 10% FBS and antibiotics in 24-well tissue plates. At near confluence, cells were treated with PB at 0, 10, 25, 50, 75 and 100 mcg/mL, in triplicate at each dose. Cells were also treated with PMA (100 ng/mL) for MMP-9 induction. Cell proliferation was assayed by MTT assay, MMP by zymography, invasion through Matrigel, apoptosis using green caspase detection, and morphology by H&E staining.

Results: PB inhibited proliferation of human fibrosarcoma cells HT-1080 by 50% at 10 mcg/mL, 60% at 25 mcg/mL and 80% at 50-100 mcg/mL, and inhibited the secretion of both MMP-2 and MMP-9 in a dose-dependent manner, with total inhibition at 50 mcg/mL concentration. Invasion through Matrigel was inhibited 100% at 25 mcg/mL PB. PB also induced apoptosis in a dose- dependent fashion. H&E staining showed slight morphological changes at higher concentrations.

Conclusion: These results suggest that PB is a potential therapeutic agent for fibrosarcoma, with potent anti-metastatic activity, because it inhibited fibrosarcoma cell proliferation, MMP-2 and -9 expression, invasion through Matrigel and apoptosis, important parameters for cancer prevention.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1500. doi:10.1158/1538-7445.AM2011-1500
Abstract 1515: Disruption of circadian rhythmicity in liver metastasis of colorectal cancer
Authors: Huisman Sander A. , van Ginhoven T.M. , van den Berg J.W. , Roest H.P. , Tamanini F. , Destici E. , IJzermans J.N.M. , van der Horst G.T.J. , de Bruin R.W.F.
Published: June 2012

Background: Liver metastases of colorectal cancer are a major cause of cancer related mortality. Resection of hepatic metastasis prolongs life, but 25% of the functional reserve of the liver should remain after resection to allow adequate liver regeneration. An effective means to downsize the tumor and enable resection is the use of chemotherapeutics. Efficacy and adverse side effects of chemotherapy may vary considerably over a 24 hour period. Chronotherapy (circadian rhythm adjusted chemotherapy) could take advantage of the fact that there may be asynchronies in circadian rhythm between normal and cancer tissue. In addition, genetic or functional disruption of the circadian clock may result in genomic instability and accelerated carcinogenisis and tumor growth.

A more thorough understanding of circadian rhytmicity may help in elucidating the role of the clock in tumorigenesis and tumor growth and develop more effective chronotherapeutic chemotherapy treatment schedules. Therefore, in this study we investigated the 24-hour expression levels of key circadian clock genes in colorectal liver metastasis and adjacent healthy liver tissue in the mouse.

Materials and methods: Male BALB/c mice were injected with C26 colorectal carcinoma cells into the spleen. Three weeks after tumor inoculation, a hepatectomy was performed at 6 different timepoints over a 24-hour period (n=4 per timepoint) to compare circadian rhythmicity in tumor and adjacent healthy tissue. RNA was isolated and RT-PCR was performed on 5 clock genes: Rev-erbα, Per2, Per1, DBP, and Bmal1 and a clock independent gene induced by feeding (Acss2).

Results: Healthy liver tissue showed normal 24-hour oscillations of all 5 clock genes, consistent with normal circadian rhytmicity. In colorectal liver metastasis however, 24-hour oscillations were completely absent in all clock genes except Per2. Per2 oscillations were similar in tumor and adjacent liver tissue, although expression levels in tumor tissue were lower. Since Per2 may also be induced by feeding, we determined the expression of the food induced gene Acss2, which is independent of circadian rhythm. Acss2 levels rose in response to feeding, followed by a rise in Per2, strongly suggesting that the increased expression of Per2 was induced by feeding.

Conclusion: Our data demonstrate the absence of circadian rhythm in hepatic metastasis of C26 coloncarcinoma in vivo. The difference between tumor and healthy tissue in this model may provide new clues to further understand the biology, and optimize treatment of hepatic metastasis.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1515. doi:10.1158/1538-7445.AM2011-1515
Abstract 1517: Retinoblastoma tumor development: Role of tumor-associated macrophages and their sub-type in LHBETATAG retinal tumor progression
Authors: Pina Yolanda , Decatur Christina , Houston Samuel , Shah Nikesh , Cavalcante Ludimila , Murray Timothy
Published: June 2012

Purpose: The purpose of the current study is to establish the distribution of tumor associated macrophages (TAMs) during tumor development in a transgenic retinoblastoma mouse model, study the contribution of bone-marrow derived macrophages in these tumors, and assess the supportive role of TAMs in retinoblastoma tumor growth.

Methods: Macrophage infiltration in transgenic retinoblastoma tumors was assessed by immunohistochemistry at different time points in tumorigenesis. To establish the origin of TAMs in transgenic retinoblastoma tumors, 107 bone marrow cells from green fluorescent protein (GFP) positive 16-week-old mice were transplanted into age-matched, irradiated LHBETATAG mice via tail vein injections. Macrophage depletion was performed by subconjuctival (SC) delivery of liposomal clodronate.

Results: TAMs’ density increased from 4- to 12-weeks of age in mice with small to medium-sized tumors (p=0.037), and remained stable in the later stages of the disease (i.e., 16-week-old with large tumors; p=0.20). 38% of TAMs (2.5 ± 3.2 cells per 400X hpf) in 16-week old mice were GFP-positive, bone marrow-derived macrophages. Total TAM depletion was correlated with a significant decrease in the expression levels of MMP-9 (p=0.014) and mature vessels (p<0.001), and a nonsignificant decrease in the density of neovessels (p=0.94). The density of M2-polarized TAMs did not change significantly following TAM depletion (p=0.68). Following M1-polarized TAM depletion, the tumor burden increased (p=0.056).

Conclusions: The current study expands our understanding of the complex role that macrophages play in retinoblastoma. Macrophage modulation in the tumor microenvironment is a critical factor in retinoblastoma tumor development.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1517. doi:10.1158/1538-7445.AM2011-1517
Abstract 1518: Gene expression profiling reveals acidosis/GPR4-induced inflammatory responses in vascular endothelial cells
Authors: Dong Lixue , Castellone Reid D. , Chi Jen-Tsan , Yang Li V.
Published: June 2012

Due to deficient blood perfusion, hypoxia, and glycolytic metabolism, acidic tissue microenvironments commonly exist in solid tumors, inflammation, and many other pathological conditions. However, the molecular mechanisms by which cells respond to acidic microenvironments are not well understood. GPR4 has recently been identified as a novel proton-sensing receptor expressed in endothelial cells, cancer cells, and other cell types. GPR4 is activated by acidic extracellular pH but has lower activation at the physiological pH 7.4 and minimal activation at more alkaline pH. To delineate the function and signaling pathways of GPR4 in endothelial cells under acidic conditions, we performed gene expression microarray analyses. Primary human umbilical vein endothelial cells (HUVEC) that express endogenous GPR4 were stably transduced with the MSCV-IRES-GFP control vector (designated as HUVEC/Vector cells) or the MSCV-GPR4-IRES-GFP construct (designated as HUVEC/GPR4 cells). HUVEC/Vector and HUVEC/GPR4 cells were treated with pH 6.4 (400 nM H+) for 5 hours to activate GPR4 or with pH 8.4 (4 nM H+) as a negative control. RNAs were isolated, labeled, and hybridized with the Agilent Whole Genome Microarray Chip to compare gene expression. The results demonstrated that the acidic pH treatment of HUVEC/Vector cells stimulated the expression of a number of inflammatory genes, which represent a predominant signature in the microarray dataset. The induction of most of the acid-responsive inflammatory genes were further enhanced by GPR4 overexpression in HUVEC/GPR4 cells at pH 6.4, suggesting these genes are responsive to varying levels of GPR4 signaling. The inflammatory genes with substantial up-regulation by acidosis/GPR4 include chemokines, cytokines, adhesion molecules, genes involved in the TNFα and NF-κB pathways, prostaglandin-endoperoxidase synthase, and EGR transcription factors. The up-regulation of the genes at pH 6.4, compared to pH 7.4 and pH 8.4, was further confirmed by real-time RT-PCR. Taken together, our results show that activation of GPR4 by acidosis stimulates the expression of a wide range of inflammatory genes in endothelial cells. Since the inflammatory response of endothelial cells is important for blood vessels to interact with immune cells and tumor cells in the microenvironment, the acidosis/GPR4 signaling may play a role in the processes such as tumor immunity and metastasis.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1518. doi:10.1158/1538-7445.AM2011-1518
Abstract 1551: Soluble tissue transglutaminase modulates cellular adhesion to matrix and peritoneal metastasis in ovarian cancer
Authors: Yakubov Bakhtiyor , Chelladurai Bhadrani , Turchi John , Matei Daniela E.
Published: June 2012

Background: Tissue transglutaminase (TG2) is a secreted protein that binds fibronectin (FN) and exerts protein transamidating activity in the presence of Ca2+. We previously reported that TG2 is upregulated in ovarian cancer (OC) cells and enhances intraperitoneal (ip) metastasis. TG2 is secreted abundantly in OC ascites, yet the function of soluble (s) TG2 remains unknown. We hypothesized that sTG2 acts as a permissive factor in the ECM by promoting cancer cell proliferation and adhesion to the matrix.

Materials and Methods: Recombinant his6-tagged full length human TG2 and its inactive enzymatic mutant C277S were purified by immobilized metal (Co2+) affinity and anion exchange chromatography. OV90 and A2780 OC cells that do not express endogenous TG2 were used. Solid phase adhesion assay measured adhesion to ECM proteins, MTT assay measured cell proliferation and an ip xenograft model quantified tumor formation and metastasis.

Results: S-TG2 enhanced cell adhesion to FN (p = 0.01) and vitronectin (VN, p = 0.01), but not to collagen and laminin. A neutralizing antibody to α5β1 and to αVβ3 blocked sTG2 induced cell adhesion to FN or to VN, respectively, suggesting that sTG2 modulates cell interaction with the matrix via integrins. Digestion of FN with matrix metallo-proteinase 2 blocked OC cell adhesion to digested fragments of FN, while addition of sTG2 restored OC cells’ binding to this protease-digested matrix (p = 0.02). This suggests that the soluble enzyme may facilitate the interaction of OC cells with a protease-remodeled matrix remodeled, similar to the tumor ECM. Enzymatically inactive soluble C277S TG2 enhanced OC cell adhesion to FN to the same extent as wild type TG2. In contrast, an antibody raised against the FN-binding domain of TG2 blocked sTG2 induced OC cell adhesion to FN (p =0.01). These data suggest that the interaction between TG2 and FN, but not the transamidating activity, is important to sTG2-mediated cell interaction with the ECM. When added to conditioned media, sTG2 (2.5ug/mL) increased OC cell proliferation by 20% (p = 0.01). Biweekly ip inoculation of 2.5ug sTG2 increased peritoneal dissemination of OV90 cells in nude mice (11.38 vs. 27.75, p = 0.02), but did not affect tumor volume (412.3 vs. 578.2 mm3, p = 0.34).

Conclusions: These data demonstrate that sTG2 increases OC cell adhesion to the ECM and peritoneal metastasis. The key mechanism involved in this process is the interaction between sTG2, FN, and integrins.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1551. doi:10.1158/1538-7445.AM2011-1551
Abstract 1557: Tumor Galectin-1 mediates lung cancer growth and metastasis through immunoregulation of T-cell apoptosis
Authors: Banh Alice , Zhang Jing , Cao Hongbin , Bouley Donna , Kwok Shirley , Kong Christina , Giaccia Amato , Koong Albert , Le Quynh-Thu
Published: June 2012

Studies have shown that Galectin-1 (Gal-1), a carbohydrate-binding protein whose secretion is enhanced under hypoxia, can promote tumor aggressiveness through its effect on angiogenesis and T-cell apoptosis. However, the importance of tumor versus host Gal-1 in tumor progression is unclear. Using matched Gal-1 wild-type and null mice together with Lewis lung carcinoma cells stably knocked-down of Gal-1 (shGal-1) or scramble control (Scr), we show that tumor Gal-1 is more critical that host Gal-1 in promoting tumor growth and spontaneous metastasis. Scr tumors had higher growth and metastasis rates compared to shGal-1 tumors, regardless of the host genotype. The enhanced growth and metastasis associated with Gal-1 appears to be related to its immunomodulatory function since it was abolished in immune-deficient mice. In contrast, angiogenesis, as assessed by microvessel density count, was similar between shGal-1 and Scr tumors when they were allowed to reach comparable size. These data indicated that tumor Gal-1 is more important than host Gal-1 in mediating tumor progression and that its effect was exerted through intratumoral T-cell modulation. These findings have broad implications in developing novel targeting strategies for Gal-1 in cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1557. doi:10.1158/1538-7445.AM2011-1557
Abstract 1565: API 31510 as a potential agent in management of CNS leukemia
Authors: Narain Niven R. , Mauro Lucia M. , Perez Paula P. , Caperton Caroline V. , Sarangarajan Rangaprasad , Jimenez Joaquin J. , McCook John P.
Published: June 2012

Infiltration of cancer cells into the cerebrospinal milieu remains a challenge to oncologists. Thus, prophylactic therapy to avoid CNS tumors is now standard of care to avoid the development of tumors in the central nervous system (CNS). Despite the advances in oncological care, this issue remains poorly understood. In this regard, we have created a model of CNS chloroleukemia using Fischer 344 rats, in which chloroleukemic cells were injected when they are newborns and lipopolysaccharide (LPS) is given as a first-line of treatment. The cure rate with this regimen is 50%, and approximately 10% of survivors develop CNS leukemia as judged by their motor skills and the presence of quadriplegia and paraplegia. For this study, we injected 2400 Fischer 344 neonates with MIAC51 and treated them with LPS. All animals with overt signs of leukemia were sacrificed by day 26. By day 35, survivors started exhibiting CNS abnormalities. Of that cohort, we selected 150 animals with hind leg paraplegia on day 40. These animals were then re-randomized into 5 groups: group 1 received no treatment, group 2 received excipient control IV, group 3 received 5mg/kg API 31510 IV, group 4 received 25mg/kg IV, and group 5 received 100 mg/kg IV for 4 weeks, 3 times daily. Rats in groups 1, 2, 3 and 4 did not exhibit any signs of improvement and were sacrificed due to metastatic malignancy. These findings were recorded by MRI positive for tumor cells. In sharp contrast, animals injected with 100mg/kg IV exhibited a significant recovery of their motor skills and regained their ability to walk. MRI distinctly shows the lack of tumor cells in this group. Taken together, these results strongly indicate that API 31510 may provide a cure for CNS leukemia and may also be an effective prophylactic agent to prevent the extravasation of leukemic cells into the CNS.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1565. doi:10.1158/1538-7445.AM2011-1565
Abstract 1570: Thermal ablation improves oxygenation in remaining viable tumor
Authors: Griffin Robert J. , Przybyla Beata D. , Pagan Jonathon D. , Chen Xin , Shafirstein Gal , Webber Jessica S. , Moros Eduardo G.
Published: June 2012

To increase the use and effectiveness of thermal therapy of various solid cancers, it is necessary to understand how tumor remaining viable after ablation responds and repairs. Tumor oxygenation and angiogenesis are critical for tumor recurrence and therefore we are studying the changes in tumor oxygenation and migration pattern of endothelial progenitor cells (EPCs), one component of angiogenesis, after local thermal ablation of tumors in mice. Tumor cells in sublethal ablation zones may release a unique set of chemokines in an attempt to attract cells to repair and regrow the tumor. The location and mechanisms by which these cells are recruited to the tumor may be distinct from other methods of thermal therapy which suggests that specific targets to improve ablation outcomes may be identified. If the oxygenation is changed in tumor that remains viable after ablation, concomitant application of chemotherapy or radiation therapy may be an effective way to improve tumor control. Murine FSaII fibrosarcoma and 4T1 breast carcinoma tumors were ablated with a conductance probe or high intensity ultrasound at an average peak temperature of 60-90C for 1.5 min. One group of mice was immediately injected IV with near infrared emitting indocyanine green (ICG)-labeled EPCs, while the other group was injected with EPCs labeled with a stable in situ dye (PKH). The mice injected with ICG-labeled EPC were optically imaged at various time points up to 72 hours. These images were then compared to those of untreated tumors to observe the migration patterns of EPC in mice that had received HIFU ablation. In other mice, pimonidazole was injected 72 h post ablation and tumors were excised and stained for pimonidazole adducts as evidence of hypoxia. We routinely ablated approximately 50% of tumor tissue, leaving the other 50% peripheral tumor tissue viable. We were able to detect an increased amount of ICG signal in the tumor that received ablation ablation, when compared to controls. Histological examination revealed the presence of EPC cells in the tumor after ablation, while the control showed no signs of EPC presence. The tumor oxygenation in viable tissue after ablation was found to increase from control tumors where 27% of the tumor was pimonidazole positive to only 5% in thermally ablated tumors. The role of endothelial progenitors and tumor oxygenation changes in overall tumor response to ablation alone and combined with other modalities is being actively studied by our group. This work was supported by NIH grant CA44114 and Central Arkansas Radiation Therapy Institute.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1570. doi:10.1158/1538-7445.AM2011-1570
Abstract 1610: Predicting NSCLC outcomes based on tumor growth in SCID mice
Authors: Hennon Mark W. , Hylander Bonnie , Morrison Carl , Miller Austin , Groman Adrienne , Dexter Elisabeth , Yendamuri Sai , Demmy Todd L. , Adjei Alex , Reid Mary , Repasky Elizabeth , Nwogu Chukwumere
Published: June 2012

Background. Xenograft models of non-small cell lung carcinoma (NSCLC) in immunodeficient mice have played an important role in the study of new chemotherapeutic regimens, including targeted agents. We hypothesize that the success of human tumor implantation in severe combined immune-deficient (SCID) mice will correlate with the observed clinical outcomes in patients with resected NSCLC. Such growth in mice may reflect the aggressiveness of human tumors.

Methods. 196 lung cancer specimens from 193 patients who underwent surgical resection of NSCLC from June 1995 to June 2010 were implanted subcutaneously in (SCID) mice. Mice were observed for a period of six months to assess the success of tumor implantation. This was determined by the growth of tumors from 2-3 mm to 10 mm in diameter. Overall and disease free survival in patients were reviewed and were correlated with tumor growth in SCID mice.

Results. Median overall and disease free survival in patients with successful tumor implantation in SCID mice was 16.3 months and 4.2 months vs. 38.6 and 20.6 months for patients whose tumor did not grow (logrank p = 0.0039 and 0.0024 respectively.) Half of all patients whose tumor successfully grew in SCID mice recurred or died within twelve months.

Conclusions. Success of tumor implantation in SCID correlates with worse clinical outcome in patients with NSCLC. This model can be expanded to evaluate the predicted response of specific tumors to novel chemotherapeutic or molecularly targeted agents.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1610. doi:10.1158/1538-7445.AM2011-1610
Abstract 1616: The OPCML tumor suppressor negatively regulates a specific repertoire of 5 receptor tyrosine kinases via a novel proteasomal mechanism, and its recombinant derivative is a potent in-vivo anticancer protein therapy
Authors: McKie Arthur B. , Vaughan Sebastian , Okon Imoh , Wong Joshua L.C. , Zanini Elisa , Lam Eric W-F , Chayen Naomi E. , Gabra Hani
Published: June 2012

OPCML, a GPI anchored tumor suppressor gene is inactivated by somatic methylation in multiple cancers. We previously identified this gene by LOH mapping and demonstrated that it was inactivated by somatic methylation in 80% of ovarian cancers. Restoring OPCML expression by stable transfection suppressed in-vitro growth and in-vivo tumorigenicity. We hypothesized that OPCML might abrogate growth signaling pathways. In SKOV-3 and PEO1, ovarian cancer cell lines with no expression of OPCML, we demonstrated that OPCML negatively regulates a specific repertoire of receptor tyrosine kinases (RTKs) EPHA2, FGFR1, FGFR3, HER2 and HER4, and reciprocally, OPCML siRNA knockdown in normal ovarian surface epithelial cells up-regulates these same RTKs, with no effect on RTKs EPHA10, FGFR2, FGFR4, EGFR, HER3, VEGFR1 and VEGFR3. Example immunoprecipitation experiments revealed that OPCML binds to EphA2, FGFR1 and HER2 extracellular domains with no such interaction to EGFR, thus OPCML binds directly to RTKS that it negatively regulates. We demonstrate that OPCML is located exclusively in the raft membrane fraction and sequesters RTKs that it binds to the raft fraction, leading to polyubiquitination and proteosomal degradation via a cav-1 endosomal mechanism resulting in systems depletion of this specific RTK repertoire, that does not occur with RTKs that OPCML does not bind. We demonstrate that OPCML abrogates EGF mediated phosphorylation of FGFR1, HER2 and EGFR and the downstream phosphosignaling of pErk and pAKT. A recombinant modified OPCML-like protein without a GPI anchor, signal peptide or glycosylation was constructed and expressed in E.Coli. This rOPCML tumor suppressor protein therapeutic caused growth inhibition by apoptosis in 6/7 ovarian cancer cell lines tested with no effect on OPCML expressing normal ovarian surface epithelium by an identical mechanism to the transfected normal protein. rOPCML was then injected intraperitoneally twice weekly in two murine intraperitoneal models of ovarian cancer (nude mouse A2780 and SKOV3) and demonstrated profound inhibition of tumour weight, ascites volume and peritoneal dissemination compared with BSA control. In Summary, the OPCML tumour suppressor mediates its suppressor function by systems level negative regulation of at least 5 RTKs and a recombinant modified derivative is a potent tumor suppressor protein therapeutic in-vitro and in-vivo.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1616. doi:10.1158/1538-7445.AM2011-1616
Abstract 1621: The proprotein convertase PCSK1 is a novel drug target in alveolar soft part sarcoma (ASPS)
Authors: Vistica David T. , Kenney Susan , Scudiero Dominic A. , Reinhart Russell A. , Selby Michael H. , Butcher Donna O. , Shoemaker Robert H.
Published: June 2012

Proprotein convertases (PCs) are a family of calcium-dependent serine endoproteases which mediate the proteolytic cleavage, at pairs of dibasic amino acids, of many precursor proteins, including growth factors and pro-hormones, into their functionally active forms. One family member, PCSK1, was identified as a potential therapeutic target in ASPS. This was based on gene expression profiling of 4 primary and 3 metastatic ASPS tumors (BMC Cancer 9:22, 2009). Affymetrix 133 Plus 2 arrays revealed that PCSK1, but not PCSK2 or furin, was significantly up-regulated (range 18.6 to 131 fold) relative to universal RNA, a multi-organ pool of non-tumor RNA's. PCSK1 expression was confirmed by immunohistochemistry and Western Blot analysis in patient tumors, in an ASPS xenograft model, and in a cell line developed from the xenograft tumor (ASPS-1; J Pediatr. Hematol. Oncol., in press 2010) as well as in tumors produced by ASPS-1 in immunocompromised mice. ASPS-1 was utilized, in-vitro, to evaluate a panel of PCSK1 siRNA's, the most potent of these produced a 75% reduction in the PCSK1 RNA transcript, reduced expression of PCSK1 protein and produced a 50% reduction in ASPS-1 growth. As part of an effort to develop new therapeutic approaches toward the treatment of ASPS we have implemented a high-throughput screening effort to identify small molecule inhibitors of PCSK1. Utilizing recombinant PCSK1 and the fluorescent substrate (Pyr-Arg-Thr-Lys-Arg-MCA) a high-throughput screen of a small molecule library (∼15 K compounds) was initiated. Results of a pilot screen of ∼2400 compounds at concentrations ranging from 0.1 to 100 uM yielded several inhibitors of PCSK1 which were retested in a 20 concentration assay to confirm activity. Preliminary analysis of structure-activity relationships identified several active inhibitor chemotypes, comparable in potency to the known peptide PCSK1 inhibitor, Ac-Leu-Leu-Arg-Val-Lys-Arg-NH2, containing aryl groups linked by aliphatic chains containing peptide bonds. Such compounds may act as substrate mimetic inhibitors and will be further evaluated for inhibition of growth of ASPS-1.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1621. doi:10.1158/1538-7445.AM2011-1621
Abstract 1627: Proteome-wide analysis of echinoderm microtubule associated protein like 4 – anaplastic lymphoma kinase (EML4-ALK) network in lung cancer
Authors: Zhang Guolin , Chen Ann , Song Lanxi , Fu Jianing , Kinose Fumi , Bai Yun , Fang Bin , Li Jiannong , Eschrich Steven , Koomen John , Haura Eric
Published: June 2012

The translocation of echinoderm microtubule associated protein like 4 – anaplastic lymphoma kinase (EML4-ALK) defines a unique, new subset of patients with non small cell lung cancer (NSCLC). It represents a novel diagnostic biomarker and therapeutic target for NSCLC. PF 02341066 is a ALK small tyrosine kinase inhibitor that has demonstrated high clinical activity in EML4-ALK- positive NSCLC patients. We sought to characterize the signaling network used by EML4-ALK and reveal potential new therapeutic targets. We used tandem affinity purification (TAP) and liquid chromatography-mass spectrometry (LC-MS/MS) to map protein-protein interactions and anti-phosphotyrosine immunoprecipitation coupled with LC-MS/MS to map tyrosine phosphorylation. We selected the human NSCLC H3122 cell line for our studies as it is sensitive to PF 02341066 and harbors ELM4-ALK. Quantitative western blotting results show that activities of MET, ALK, AKT, ERK and STAT3 were inhibited by treatment with PF 02341066. Proteome-wide analysis of tyrosine phosphorylation of the ALK signaling pathway were examined. In total, 311 tyrosine phosphorylated proteins (487 unique phosphorylated sites) were identified. Of those, 73 pTyr sites (62proteins) were upregulated and 78 pTyr sites (58 proteins) were downregulated by 1uM PF02341066. Using TAP, we identified 13 proteints interacting with EML4-ALK including GRB2 and SHC1. Core interactions were verified by coupling of immunoprecipitation to Western blot and mass spectrometry. A comprehensive physical ALK network was generated by integrating both phosphoproteomic data and interactional partners using pathway analysis algorithms. Two adaptors, SHC1and GAB2, are suggested as the first outlets of signaling propagation from ALK. Other sublevels directly connected with ALK may allow further understanding the mechanism of ‘ALK addiction’. A sub-network composed of 35 cell death associated proteins was created. An ongoing functional analysis including siRNA and small inhibitor screen on this small network should validate druggable targets. This network approach has potential translational applications to identify novel approaches to treatment of ALK addicted lung cancers and could be useful in patients with acquired resistance to ALK inhibitors.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1627. doi:10.1158/1538-7445.AM2011-1627
Abstract 163: mRNA and miRNA profiling of platinum-resistant non-small cell lung cancer cell clones identifies potential targets involved in DNA metabolism and DNA replication
Authors: Salim Hogir M. , Lundholm Lovisa , Zong Dali , Hååg Petra , Mörk Birgitta , Viktorsson Kristina , Lewensohn Rolf
Published: June 2012

Non small cell lung cancer (NSCLC) constitutes the majority of lung cancer diagnoses and platinum-based chemotherapy regimen is the standard treatment. Unfortunately, a large proportion do not respond or relapse on such treatment regimens for yet only partly revealed mechanisms. We profiled residual clones of NSCLC cells after treatment with cisplatin. The mRNA and miRNA pattern was compared between untreated and cisplatin-resistant NSCLC clones using Affymetrix gene and miRNA expression arrays, respectively.

We found that around 1100 genes were up regulated (at least two fold) and about 1200 genes were down regulated (at least two fold) in the cisplatin-resistant clones compared with untreated NSCLCs. We then used the Gene Set Enrichment Analysis (GSEA) to find the over- and underrepresented gene sets in large datasets like, Gene Trail and Gene Ontology. We found that cisplatin-refractory NSCLCs showed increased expression of genes involved in different cellular mechanisms including DNA metabolic processes, DNA replication and DNA-dependent DNA replication initiation for example, XRCC2, FANCI, FANCD2, RRM1, MRE11A, BLM, RRM2 and TOP2A which were validated on RNA, protein and functional level for their role in cisplatin sensitivity of NSCLC.

We also found a few miRNAs that were up regulated in cisplatin refractory NSCLC by a least 1.3-fold e.g. miRNA-25 star and miRNA-1290. Interestingly, only 14 miRNAs were down regulated (by at least 1.3 fold) in the surviving cisplatin refractory NSCLC cells. Among these miRNAs were (miR-29b, miR-210, miR-486-3p, miR-324-5p, miR-572, miR-30a, e, miR-424, miR-449a, b, miR-301a, miR-153, miR-150 star and miR-34c-5p). We then looked for the predicted targets of these miRNAs using Target scan and microcosm and we found a large list of genes to be potential targets of the miRNAs among them are genes involved in apoptosis, cell cycle regulation, Wnt signaling pathway and other cellular processes (ex. VDAC, SMAD2, RAN, etc.). Correlation analysis between miRNA gene targets and gene expression data is ongoing with the aim to identify and validate putative targets on RNA, protein and functional level.

In conclusion, our data show that cisplatin refractory NSCLC tumors have dysregulated mRNAs and miRNAs. Among the dysregulated genes, are certain genes involved in DNA metabolism, DNA replication and DNA-dependent DNA replication initiation which may be novel targets for improved platinum-based therapy response of NSCLC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 163. doi:10.1158/1538-7445.AM2011-163
Abstract 1636: ErbB4 is a new therapeutic target for metastasis and anoikis resistance in Ewing's sarcoma
Authors: Mendoza-Naranjo Ariadna , Mistry Pitri , Lazic Nikola , Strauss Sandra J. , Sorensen Poul HB
Published: June 2012

Most Ewing sarcoma (ES)-related deaths occur as a result of metastatic disease. As a barrier to metastases, cells normally undergo a form of cell death called ‘‘anoikis” after they lose contact with the extracellular matrix or neighboring cells. Tumor cells that acquire malignant potential develop mechanisms to suppress anoikis. We previously demonstrated that ES tumors suppress anoikis and show prolonged survival as multicellular spheroids, which mimic micrometastases under anchorage-independent conditions. Here we have used the paired CHLA-9 primary tumor line established at diagnosis and the metastatic CHLA-10 line established from the same patient after four cycles of induction chemotherapy with cisplatin, doxorubicin, cyclophosphamide, and etoposide, as well as high-throughput Affymetrix whole exon array platforms to screen for potential therapeutic targets to treat metastatic disease. We have identified ErbB4, a member of the epidermal growth factor receptor (EGFR) family, as a novel mediator of anoikis resistance in ES. We detected a 4-fold increase in ERBB4, along with enhanced ErbB4 protein expression and Tyr kinase activation (confirmed by P-Tyr kinase proteome arrays) in CHLA-10 cells grown under anchorage independent conditions, compared to CHLA-9 tumor cells under the same conditions. Upregulated ErbB4 expression was further confirmed in a panel of metastatic vs. primary ES cell lines at both the mRNA and protein levels. We are currently validating these findings in the clinical setting by analyzing ErbB4 expression in paired primary vs. metastatic biopsies from ES patients. shRNA-mediated knockdown of ErbB4, or treatment with the ErbB inhibitor Lapatinib, dramatically reduced proliferation, decelerated tumor invasion, and sensitized metastatic/chemoresistant ES tumors to chemotherapeutic drugs used to treat ES patients. Taken together, our findings identify ErbB4 as a novel target for metastasis and anoikis resistance in ES.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1636. doi:10.1158/1538-7445.AM2011-1636
Abstract 1657: A pivotal role for epithelial cell adhesion molecule (EpCAM) in survival of lung cancer cells
Authors: Hase Tetsunari , Sato Mitsuo , Yoshida Kenya , Takeyama Yoshihiro , Elshazley Momen , Sekido Yoshitaka , Gazdar Adi F. , Minna John D. , Kondo Masashi , Hasegawa Yoshinori
Published: June 2012

Introduction: Epithelial cell adhesion molecule (EpCAM) is overexpressed in a wide variety of human cancers, including lung cancer, and its contribution to increased proliferation through up-regulating cell cycle accelerators such as cyclin A/E is well demonstrated in breast and gastric cancers. Nevertheless, very little is known about its role in survival of cancer cells. In addition, functional role of EpCAM in the pathogenesis of lung cancer remains to be explored.

Matrials and methods: 18 human non-small cell lung cancer cell lines and an immortalized normal human bronchial epithelial cell (HBEC4) lines were used. EpCAM expression was measured by quantitative real-time PCR (qRT-PCR) and FACS. RNA interference (RNAi)-mediated gene silencing for EpCAM was done in three non-small cell lung cancer cell and HBEC4 lines. Cell proliferation was measured by WST-1 and clonogenic growth was measured by liquid and soft agar colony formation assays. Apoptosis was evaluated by annexin V/7-AAD staining. FACS with PI staining was done to examine apoptosis and cell cycle. p27kip1 expression was evaluated by western blot analysis. In vitro invasion assay was done by using transwell chambers layered with matrigel and invading cells were stained and counted.

Results: We analyzed the expression of EpCAM in a panel of lung cancer cell and HBEC4 lines and found that most of lung cancer cell lines expressed EpCAM. EpCAM knockdown suppressed proliferation and clonogenic growth of two EpCAM-expressing lung cancer cell lines in anchorage dependent and independent conditions. EpCAM knockdown suppressed invasiveness in a highly invasive line but not in a lowly invasive cell line. In addition, EpCAM knockdown induced massive apoptosis in both cell lines as well as another EpCAM-expressing lung cancer cell line but to a much lesser extent in HBEC4 line. EpCAM knockdown caused lung cancer cells to have increased responsiveness to contact inhibition in part through up-regulating p27kip1 cyclin dependent kinase inhibitor. Conclusion: These results demonstrate that EpCAM substantially contributes to the pathogenesis of lung cancer, especially in its survival, and the development of EpCAM-targeted therapy for lung cancer may have promise.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1657. doi:10.1158/1538-7445.AM2011-1657
Abstract 1675: Identification of KURO3 overexpressed in high-grade prostate cancer
Authors: Tamura Kenji , Furihata Mutsuo , Satake Hirofumi , Anchi Takashi , Ashida Shingo , Yamasaki Ichiro , Inoue Keiji , Shuin Taro
Published: June 2012

Prostate cancer is usually androgen-dependent and responds well to androgen ablation therapy. However, at a certain stage some prostate cancers eventually acquire a castration-resistant phenotype. These cancer cells are originally high-grade and show very poor response to any anticancer therapies. To characterize the molecular features of clinical high-grade prostate cancers, we analyzed gene-expression profiles by genome-wide cDNA microarray combined with microdissection and found dozens of trans-activated genes in clinical high-grade prostate cancers. Among them, we report the identification of a new biomarker, KURO3. We confirmed overexpression of KURO3 in high-grade prostate cancer cells by RT-PCR. Knockdown of KURO3 expression by siRNA in a prostate cancer cell line resulted in a drastic decrease of cell numbers. In contrast, KURO3 overexpression in a prostate cancer cell line promoted cell proliferation. These findings suggest that KURO3 could be involved in aggressive phenotype of prostate cancers, including hormone-resistant prostate cancers, and that it should be a potential molecular target for development of new therapeutics and a diagnostic biomarker for aggressive prostate cancers.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1675. doi:10.1158/1538-7445.AM2011-1675
Abstract 1679: Increased expression of hCAS/CSE1L in colorectal cancer: Correlation with tumor progression
Authors: Henderson-Jackson Evita , Eschrich Steven A. , Chen Dung-Tsa , Yeatman Timothy J. , Coppola Domenico , Malafa Mokenge P.
Published: June 2012

Introduction. The human cellular apoptosis susceptibility (hCAS) gene product has been shown to be involved in cellular proliferation, apoptosis, invasion and metastasis. CSE1 is a yeast gene with high homology to hCAS (CSE1-like or CSE1L) which has been shown to be involved in chromosome segregation during mitosis. hCAS has also been shown to be a key player in cytoplasm-to-nuclear transport by shuttling importin-alpha from the nucleus back out into the cytoplasm. We hypothesized that the expression of hCAS may be altered during the progression of human colorectal cancer (CRC).

Methods. In order to examine hCAS at the gene expression level, we surveyed three different datasets from studies in which Affymetrix U133+gene chips had been performed on patients with CRC, as well as patients with adenomas and normal individuals. We examined relative gene expression levels for 3 probe sets on the U133+ gene chip that closely align to the refseq for the hCAS gene. hCAS protein expression level was semiquantitatively measured using immunohistochemistry (IHC) and the tissue microarray (TMA) technique. A CRC TMA had been previously created at the Moffitt Cancer Center using resection specimens of primary CRC of different stages, and inclusive of adenomas and normal tissues samples from the same patients.

Results. hCAS gene expression was increased in all adenoma and CRC samples tested by microarray, compared to histologically normal colonic mucosa. However, the difference in hCAS mRNA levels between adenoma and CRC samples was not statistically different. No correlation was found between hCAS and cancer stage, grade, vascular invasion, sex and/or age. Interestingly, in the invasive CRCs, intense hCAS proteins expression was localized to both nucleus and cytoplasm. Conversely, early stage lesions showed predominantly cytoplasmic hCAS staining.

Conclusions. Our results demonstrate that hCAS protein is expressed early and across all stages of CRC development. Because of its early expression hCAS may represent a marker for early detection of CRC. In addition, because of its involvement in a variety of important cellular processes, this gene product might represent a new potential target for prevention and treatment of CRC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1679. doi:10.1158/1538-7445.AM2011-1679
Abstract 1684: 2-Methoxyestradiol reverses the epithelial-mesenchymal transition in Barrett's adenocarcinoma cells
Authors: Chattopadhyay Indranil , Majumder Monami , Haque Inamul , De Archana , Wickliffe Joi , Banerjee Snigdha , Van Veldhuizen Peter J. , Banerjee Sushanta K. , Kambhampati Suman
Published: June 2012

2-Methoxyestradiol (2-ME2) is a novel anti-cancer agent because of its ability to potentiate apoptotic cell death and inhibit cancer cell growth and angiogenesis. In our study, we have investigated whether 2-ME2 is able to modulate epithelial-mesenchymal transition (EMT) in Barrett's esophageal adenocarcinoma cell (OE33), which has been postulated as an absolute requirement for tumor invasion and metastasis. Expression of epithelial (E-cadherin, β-catenin, Keratin-19) and mesenchymal markers (Oct-4, Vimentin) and stem cell markers (CD44, CD24) were studied at mRNA and protein levels in OE33 cells that were treated with 5µmolar/L of 2-ME2 for 24 h and 48 h. Our results indicate expression of β-catenin and E-cadherin is activated in a 2-ME2 dependent fashion in OE33 cells. Expression of Vimentin and Oct-4 is significantly down-regulated in 2-ME2 treated OE33 cells. Altogether, these results provide new insights into the role of 2-ME2 in EMT modulation, which may facilitate development of new therapeutic strategies to prevent the tumor invasion and metastasis.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1684. doi:10.1158/1538-7445.AM2011-1684