bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 644: A novel Gemini vitamin D analogue represses the expression of a stem cell marker CD44 in breast cancer
Authors: So Jae Young , Lee Hong Jin , Smolarek Amanda K. , Paul Shiby , Maehr Hubert , Uskokovic Milan , Cai Li , Liu Fang , Suh Nanjoo
Published: February 2011

Breast cancer stem cells are defined as cancer cells with self-renewal capacity. Cancer stem cells are resistant to current chemotherapy and radiation, making it difficult to investigate the mechanisms of resistance and the therapeutic targets of the tumor initiating cell population. Recently, CD44 has been recognized as a useful cell-surface marker for detecting breast cancer stem cells, and the CD44+/CD24- phenotype selects a population with stem cell properties similar to basal-like cancer cells. CD44 is a multi-functional transmembrane protein involved in cell proliferation, angiogenesis, invasion and metastasis. The CD44 positive subpopulation of breast cancer cells are enriched in residual breast cancer cell populations after conventional therapies, suggesting that CD44 positive cells may be responsible for the recurrence of cancer and therefore it may be an important therapeutic target. We have previously shown that novel Gemini vitamin D analogs inhibit the proliferation of breast cancer epithelial cells and suppress tumorigenesis in different animal models of breast cancer. Since therapies targeting CD44 by vitamin D have not been investigated, we examined whether a novel Gemini vitamin D analog regulates CD44 expression in cultured human breast cancer cells or in mammary tumors. In basal-like MCF10DCIS.com human breast cancer cells, the Gemini vitamin D analog BXL0124 significantly inhibited the expression of CD44 protein and reduced the transcriptional activity of the CD44 promoter in vitro. To evaluate the inhibitory effects of Gemini vitamin D on CD44 expression in vivo, MCF10DCIS.com cells were xenografted into immunodeficient mice. When administered intraperitoneally (0.1 microgram/kg body weight) or orally (0.3 microgram/kg body weight) once a day for 6 weeks, Gemini vitamin D BXL0124 inhibited tumor growth and markedly decreased the expression of CD44 in the xenograft tumors. To determine whether the repression of CD44 is dependent on the vitamin D receptor (VDR), siRNA VDR was employed. The repression of CD44 expression by BXL0124 was significantly blocked by siRNA VDR in MCF10DCIS.com cells, indicating that the regulation of CD44 expression by BXL0124 is a VDR-dependent event. Taken together, the novel Gemini vitamin D analog inhibits CD44 expression in MCF10DCIS.com cells in vitro and in xenograft tumors, suggesting an inhibitory role of Gemini vitamin D analogs on cancer stem cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 644.
Abstract 659: Overexpression of TPX2 accelerates cell cycle progression in melanoma and is an indicator of response to Aurora A kinase inhibitors
Authors: Li Bin , Palma M. Dalla , Nathanson K. L. , Xu G. , Smalley K. , Herlyn M.
Published: February 2011

There is a tremendous need to identify more effective treatments for metastatic melanoma. In order to detect genetic changes in melanoma that might facilitate the development of novel therapies, we performed aCGH in melanoma cell lines and tumors and identified that Targeting Protein for Xklp2 (TPX2) was the most commonly amplified gene on 20q11.21, a region of frequent gain. Gene expression profiling studies also demonstrated overexpression of TPX2, suggesting a previously unknown role in the development of melanoma. TPX2 is a microtubule-associated protein that regulates bipolar spindle assembly and centrosome replication; the TPX2 - Aurora A interaction is essential for accurate segregation of chromosome and mitosis progression. We evaluated a potential role for TPX2 overexpression in melanoma using TPX2 overexpressing melanoma cell lines as a model. TPX2 overexpressing cell lines display accelerated cell cycle progression, increased CDK1/CDK2 activity, cyclin A/B1 levels, and constitutive activation and overexpression of Aurora A. We depleted TPX2 using shRNA to further dissect its role in melanoma. In unsynchronized cells, TPX2 depletion causes prolonged mitotic arrest accompanied by persistent activation of the spindle-assembly checkpoint (SAC) and caspase-3, and subsequent caspase-dependent cell death. In contrast, with G1/S synchronization, TPX2 depletion decreases S phase entry and delays S-G2-M phase transition followed by marked cell death. Accordingly, TPX2 depletion was associated with decreased CDK1 activity but not CDK2 activity, and activation of caspase and low activation of the SAC. Our data demonstrate that TPX2 overexpression is required for increased CDK1-cyclinB1 activity, thus accelerating cell cycle progression. Accordingly, TPX2-overexpressing melanoma cell lines are dependent on TPX2 for proliferation, whereas non-overexpressing cell lines and melanocytes are not. Additionally, TPX2-overexpressing cell lines are 3-10 time more sensitive to an Aurora A kinase inhibitor (VX680) than non-overexpressing cell line, suggesting that TPX2-activating Aurora A kinase could become a novel therapeutic target for treatment of melanoma with highly expressed TPX2, Finally, TPX2-IHC staining demonstrated that a significantly increased expression of TPX2 in primary and metastatic melanoma as compared to begin nevi, suggesting it is a marker of progression. Thus, we have identified TPX2 as a novel gene upregulated in melanoma. Overexpression of TPX2 may be a marker of tumors sensitive to Aurora A inhibitors allowing patient stratification for optimal therapeutic selection.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 659.
Abstract 662: Molecular characterisation of ERG, ETV-1 and PTEN- gene loci identifies patients at low and high risk of death from prostate cancer
Authors: Reid Alison , Attard Gerhardt , Ambroisine Laurence , Fisher Gabrielle , Kovacs Gyula , Brewer Daniel , Clark Jeremy , Flohr Penny , Edwards Sandra , Berney Daniel M. , Foster Chris S. , Fletcher Anne , Gerald William L. , Moller Henrik , Reuter Victor E. , Scardino Peter T. , Cuzick Jack , De Bono Johann S. , Cooper Colin S.
Published: February 2011

Background: The discovery of ERG/ETV-1-gene rearrangements and PTEN-gene loss suggests their use in a mechanism-based prognostic classification of prostate cancer (PCa).

Purpose: To evaluate the potential clinical significance and natural history of different disease categories by combining ERG/ETV-1-gene rearrangements and PTEN-gene loss status.

Methods: We utilized fluorescence in situ hybridization (FISH) assays to detect PTEN-gene loss and ERG/ETV-1-gene rearrangements in 308 conservatively managed PCa patients with survival outcome data.

Results: ERG/ETV-1-gene rearrangements alone and PTEN-gene loss alone each failed to show a link to survival in multivariate analyses. However, there was a strong interaction between ERG/ETV-1-gene rearrangements and PTEN-gene loss (p<0.001). The largest subgroup of patients (54%), lacking both PTEN-gene loss and ERG/ETV-1-gene rearrangements comprised a ‘good prognosis’ population exhibiting favourable cancer-specific survival (85.5% alive at 11 years). The presence of PTEN-gene loss in the absence of ERG/ETV-1-gene rearrangements identified a patient population (6%) with poorer cancer-specific survival that was highly significant (HR = 4.87, p<0.001 in multivariate analysis, 13.7% survival at 11 years) when compared to the ‘good prognosis’ group. ERG/ETV-1-gene rearrangement and PTEN-gene loss status should now prospectively be incorporated into a predictive model to establish whether predictive performance is improved.

Conclusions: Our data suggest that FISH studies of PTEN-gene loss and ERG/ETV-1-gene rearrangements could be pursued for patient stratification, selection and hypothesis-generating sub-group analyses in future PCa clinical trials and potentially in patient management.

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

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 662.
Abstract 663: The comparative efficacy of the PARP inhibitors AG14361, ABT888 and AZD2281 in the treatment of triple negative and BRCA1 breast cancer
Authors: Velikodvorskaya Tatiana , Fatima Naheed , Fountain Renee Quick , De Soto Joseph A.
Published: February 2011

Introduction: Most preclinical studies have used “in vitro” animal cell lines, “in vivo” allografts and/or gene knockdowns to predict the clinical efficacy of PARP1 and PARP1 & 2 inhibitors in the treatment of human breast cancer. It has been shown however that “in vitro” experiments with human breast cancer cell lines are more useful in predicting the clinical performance of cancer drugs than these other methods. In this study, the PARP1 inhibitor AG14361 and the PARP1 & 2 inhibitors AZD2281 and ABT888 were tested against a panel of 25 human breast cancer cell lines. Methods: The MTT assay was used to determine the IC50 inhibitory effect of the PARP inhibitors against the human breast cancer cell lines over a 72 hour period with each experiment being replicated at least 10 times. The MTT assay was validated for each cell line using direct counting via hemacytometer & trypan blue staining. Human to mouse xenografts were then challenged by PARP inhibitors. The IC50 results were then correlated with the PARP activity of each cell line. Results: The PARP1 inhibitor AG14361 was 5 and 8 fold more effective in inhibiting human breast cancer growth than ABT888 and AZD2281 respectively. The average IC50 values for AG14361, ABT888 and AZD2281 against BRCA1-/- human breast cancer cell lines were 20, 120, and 190 micromolar respectively. The average IC50 values for non-BRCA1 triple negative breast cancer were 24, 110 and 165 micromolar respectively. ANOVA analysis sustained the null hypothesis between BRCA1-/- breast cancer and non-BRCA1 triple negative breast cancer. Xenograft studies mirrored the “in vitro results”. These IC50 values are significantly higher than previously published preclinical data that utilized BRCA1 -/- animal cell lines and human BRCA1 gene knockdowns. This suggests that BRCA1 animal cell lines are more sensitive to PARP1 inhibitors and that human BRCA1 -/- gene knockdowns may be fundamentally different from naturally derived BRCA1-/- breast cancer cell lines. Conclusion: The PARP1 inhibitor was more effective than the PARP1 & 2 inhibitors in inhibiting breast cancer growth. Non-BRCA1 triple negative breast cancers were at least as sensitive to PARP inhibition as BRCA1 breast cancers.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 663.
Abstract 668: Synergistic Bcr/Abl degradation effect is triggered by MB12066 with NQO2 inhibitors in chronic myelogenous leukemia
Authors: Kwon Hyun-Joo , Heo Jun-Young , Park Ji-Hoon , Seo Kang-Sik , Shin So-Yeon , Jing Kaipeng , Song Kyoung-Sub , Park Jong-il , Yoon Wan-Hee , Lim Kyu , Hwang Byung-Doo , Kweon Gi-Ryang
Published: February 2011

Mutations of Bcr-Abl protein reduce the sensitivity of imatinib treatment for chronic myelogenous leukemia (CML). The resistance against imatinib treatment to CML patients needs to develop another target drug of Bcr-Abl through a novel mechanism. In present study, we investigated the effect of MB12066, small molecule related to quinone derivatives, on k562 cells (CML cell line) and the expression level of Bcr-Abl protein down regulated by MB12066. When k562cells were treated with MB12066, dose-and time-dependent Bcr-Abl degradation was detected, followed by decreasing of procaspase-3 and cleavage of PARP, which represent apoptotic stimuli. Quinone oxidoreductases [NAD(P)H: quinone Oxidoreductase 1(NQO1) and NRH: quinone oxidoreductase2 (NQO2)] are the major enzymes involved in the bioreduction of quinone-containing drugs. We found that MB12066 was a substrate of both NQO1 and NQO2 by examining recycling assay. For the functional study of NQO1 and NQO2 in Bcr-Abl degradation, we used inhibitors for each enzymes, dicumarol, quercetin and imatinib respectively. Interestingly, Bcr-Abl degradation was inhibited by dicumarol, but it was accelerated by quercetin and imatinib. Collectively, these results demonstrate that MB12066 destruct the Bcr-Abl through the NQO1 and NQO2 activity and it may show the possibility that combination therapy with NQO2 inhibitor may be beneficial for treating CML patients.

* H.J. Kwon and J.Y. Heo contributed equally to this work.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 668.
Abstract 67: Selective cytotoxic targeting of von Hippel-Lindau-deficient renal cell carcinoma cells
Authors: Lai Edwin W. , Chan Denise A. , Hay Michael P. , Giaccia Amato J.
Published: February 2011

Loss of the von Hippel-Lindau (VHL) tumor suppressor gene, an essential alteration in the development of renal cancer, can be targeted through a synthetic lethal manner. In our screen we identified a compound, STF-31536, which is selectively cytotoxic toward VHL-deficient renal cancer cells in a HIF-dependent manner. This compound did not induce apoptosis and inhibits glucose transport. These findings are significant since we have previously shown another compound STF-62247 was cytotoxic toward VHL-deficient cells and induced autophagy in a HIF-independent manner. We have also shown that STF-31536 induces cell death in vitro and in vivo, however the mechanism and target of this compound is currently unknown. In order to identify the target of STF-31536, we have coupled analogs of STF-31536 to an affinity resin. Activity of these analogs was confirmed by cytotoxicity assays. Selected affinity chromatography reagents coupled to resin were used for binding assays to determine direct drug-target interactions. The potential targets were purified and identified by gel electrophoresis after collecting fractions from the column. Renal cell carcinoma cells deficient in VHL are characterized by elevated expression of VEGF and glucose transporter 1 (Glut-1). Immunoblots for Glut-1 transporter in protein fractions eluted from the column show binding of STF-31536 to Glut-1. Therefore, we show loss of the von Hippel-Lindau (VHL) tumor suppressor gene can be targeted by this small molecule through a synthetic lethal manner by disrupting glucose transport.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 67.
Abstract 670: Aurora kinase inhibitor VE 465 synergistically enhances cytotoxicity of carboplatin in ovarian cancer cells through induction of apoptosis and downregulation of histone 3
Authors: Fu Siqing , Li Yangfan , Shen De-yu , Zheng Hong , Kavanagh John J. , Bast Rebert C. , Sood Anil K. , Hu Wei
Published: February 2011

Aurora kinases are essential for the regulation of chromosome segregation and cytokinesis during mitosis and play a role in tumorigenesis and progression in human tumors, including ovarian cancer. Aurora A and Aurora B are frequently overexpressed in high-grade and low-grade ovarian cancers. Targeting aurora kinases has great potential for improving the efficacy of treatment of ovarian cancer. In this study, we investigated whether the Aurora kinase inhibitor VE 465 will enhance the antitumor activity of carboplatin in human ovarian cancer cells. Cell viability was tested using the MTT assay in seven established human epithelial ovarian cancer cell lines with varying p53 status. We found that VE 465 and carboplatin had a synergistic effect (defined by the combination index) in both platinum-sensitive and -resistant ovarian cancers and that the growth-inhibitory effect was accompanied by reduction in expression of histone 3 and an increase in apoptosis. Aurora kinase inhibition in combination with carboplatin chemotherapy is an attractive combination for further development.

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

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 670.
Abstract 671: Triggering of CD99 induces apoptosis of Ewing's sarcoma cells through activation of p53 functions
Authors: Guerzoni Clara , Terracciano Mario , Manara Maria Cristina , Picci Piero , Scotlandi Katia
Published: February 2011

CD99 is a transmembrane 32 KDa protein, whose expression is constantly associated to Ewing's Sarcomas Family of Tumors (EWSFT), a class of child bone tumors with particular poor prognosis. The monoclonal antibody 0662, raised against CD99, triggers massive and rapid cell death of Ewing Sarcoma's cells in vitro and in vivo, through a non-canonical, non caspase-dependent apoptosis. To elucidate the underlying molecular mechanism, we evaluated by phosphor- and micro-arrays specific pathways triggered by CD99 engagement in 6647 Ewing Sarcoma cell line. Despite its role in physiological and cancer scenery has not been yet fully elucidated, CD99 appears to be sufficient to modulate many important processes involved in: I) adhesion; II) migration and metastasis; III) cell-cycle regulation and cell signaling; IV) apoptosis and death receptor signaling. These data are consistent with our previous analysis of 0662 effects in vivo and in vitro: anti-CD99 monoclonal antibody triggers, indeed, homotypic aggregation and a rapid death response of ES cells, thus resulting in growth inhibition and reduced metastatic/colony-forming potential. As pointed out by array studies and subsequent western blotting validations, upon 0662 treatment, the oncosuppressor protein p53 appears to be readily activated by phosphorilation on serine 15. We therefore evaluated p53 and MDM2 activation and sub-cellular localization upon CD99 engagement both in p53 WT (LAP35 and WE68) or mutated (6647 and TC-71) cell lines: WT (LAP35 and WE68) or mis-sense point mutated p53 (6647: S241F) cell lines display a higher sensitivity to 0662 if compared to p53 truncated cell lines (TC71). Consistently we demonstrated that whereas 0662 readily induces p53 canonical targets (p21, BAX) in p53 expressing cell lines, no increase in p21 levels is detectable in TC71 p53TRUNC cells. We also demonstrated that, upon 0662 treatment, decreased or delocalized MDM2 levels may account both for p53 activation and sustained IGF-IR signaling, since both p53 and IGF1-Rß are targeted by the ubiquitine ligase for degradation. Since most Ewing Sarcoma patients display unmodified p53 status, its functional inactivation -e.g. through MDM2/MDM4 overexpression- may account for gained tumor aggressiveness, or progressed disease. Nonetheless, involvement of TP53 might give reason for higher effectiveness we reported against local tumors and metastases in the combined employment of CD99 monoclonal antibodies with Doxorubicin and may support 0662 mAb treatment in association with other drugs targeting e.g. the p53 regulators MDM2-MDM4. (grants from EU project Eurobonet and Italian Association for Cancer Research)

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 671.
Abstract 677: Targeting the survivin splice variant 2β in taxane-resistant ovarian cancer
Authors: Vivas Pablo E. , Rodríguez-Aguayo Cristian , Han Hee-Dong , Shahzad Mian M. , Sood Anil K. , Lopez-Berestein Gabriel
Published: February 2011

Survivin, a protein highly expressed in human cancers is involved in both control of cell cycle progression and inhibition of apoptosis. Transcription of the survivin gene locus (BIRC-5) yields at least 5 alternative splice variants that have been designed survivn-WT, survivin-ΔEx3, survivin-2β, surivivn-3β, and survivin 2α. It has been shown that the survivin splice variants have different expression levels and subcellular localization patterns that could be associated with unique functional properties. Whereas the effects of survivin on apoptosis and mitosis have been extensively studied and explored therapeutically, the role of specific survivin splice variants in ovarian cancer has not been investigated. Our compelling studies indicate that, compared with their taxane-sensitive counterparts, taxane-resistant ovarian cancer cells express higher survivin mRNA (particularly the WT, ΔEx3 and 2β isoforms) levels. Studies have shown that survivin-2β has proapoptotic properties: whereas, forced expression of survivin-2β sensitized cells to taxane-induced apoptosis, small-interference RNA (siRNA)-based silencing of survivin-2β protected cells from cells from apoptotic cell death. We have observed opposite results: siRNA-based silencing of survivin-2β induced cell growth arrest and apoptosis of taxane-resistant ovarian cancer cells. Furthermore, liposomes-encapsulated siRNA targeting survivin-2β inhibited tumor growth in orthotopic murine models of ovarian cancer. The anti-tumor effect was further increased by combination with docetaxel. Finally, we found a significant association of survivin-2β expression with progression free survival in the 117 epithelial ovarian cancers obtained at primary debulking therapy. Further studies are necessary to clarify the role of survivin-2β in ovarian cancer. The key finding from our study is that survive-2β is present in a substantial proportion of ovarian cancers, and is predictive of decreased progression-free survival (PFS) of ovarian cancer patients. These studies also show the feasibility of survivin-2β-targeting siRNA liposomal as a clinically applicable therapeutic modality for taxane-resistant ovarian cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 677.
Abstract 680: Quinacrine sensitizes hepatocellular carcinoma cells to TRAIL via up-regulating DR5 and eradicating MCL-1
Authors: Wang Wenge , Katz Sharyn I. , Abdulghani Junaid , Dicker David , El-Deiry Wafik S.
Published: February 2011

Quinacrine has been widely explored in treatment of malaria, giardiasis, and rheumatic diseases. We find that quinacrine stabilizes p53 and induces p53-dependent and independent tumor cell death. Structurally, quinacrine bears similar characteristics with DNA-intercalating agents. Indeed, quinacrine competes with Propidium Iodide and shows a high binding affinity to DNA. Interestingly, quinacrine is a fluorescent dye that can be visualized under a laser microscope. In live cells, quinacrine is mainly clustered in the cytoplasm with a faint fluorescent signal in the nucleus. If the cells are fixed with methanol, quinacrine accumulates predominantly in the nucleus. Treatment by quinacrine alone at concentrations of 10-20 μM for 1-2 days can not kill hepatocellular carcinoma cells, such as HepG2, Hep3B, Huh7, which are also resistant to TRAIL. However, quinacrine renders these cells sensitive to treatment of TRAIL significantly. Co-treatment of these cells with quinacrine and TRAIL induces overwhelming cell death in 3-4 hr. Western blotting shows the level of DR5, a pro-apoptotic death receptor of TRAIL, is increased while MCL-1, a member of the Bcl-2 family, is decreased. While the synergistic effect of quinacrine with TRAIL appears to be in part independent of p53, knockdown of p53 in HepG2 cells by siRNA results in more cell death after treatment by quinacrine and TRAIL. The mechanism of quinacrine sensitization of hepatocellular carcinoma cells and the potential for clinical application are being further explored.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 680.
Abstract 681: Knockdown of connective tissue growth factor and treatment with temozolomide inhibited invasion, angiogenesis, and tumorigenesis of human glioblastomas
Authors: George Joseph , Banik Naren L. , Ray Swapan K.
Published: February 2011

Connective tissue growth factor (CTGF) is a putative proto-oncogene and plays a crucial role in endothelial cell migration and tumor angiogenesis. CTGF is highly upregulated in proliferating endothelial cells and glioblastoma cells. In this investigation, we examined whether knockdown of CTGF at the mRNA level and treatment with temozolomide (TMZ) could inhibit cell invasion, angiogenesis, and growth of human glioblastoma SNB-19 and T98G cells in vivo. The cells were stably transfected with a plasmid vector carrying the human CTGF siRNA cDNA and then treated with 10 μM TMZ for 48 h. Semiquantitative PCR, Western blotting, and immunohistochemical staining demonstrated 80% downregulation of CTGF both at mRNA and protein levels after stable transfection. Matrigel invasion, cell migration from spheroids, and cell proliferation studies demonstrated significant inhibition of cell invasion, migration, and proliferation, respectively, of both cell lines after downregulation of CTGF and treatment with TMZ. In vitro and in vivo angiogenesis assays demonstrated inhibition of network formation of endothelial cells and neovascularization under the dorsal skin of nude mice, respectively, after knockdown of CTGF and treatment with TMZ. Both subcutaneous and intracerebral tumorigenesis in nude mice was markedly reduced in CTGF downregulated cells after TMZ treatment. Mechanistic studies demonstrated significant reduction of PCNA, VEGF, c-Myc, CDK2, CDK4, and cyclin D1 and upregulation of the cell cycle inhibitors p21Waf1 and p27Kip1. Flow cytometric analysis showed cell cycle arrest at G2/M phase in both cell lines after CTGF knockdown and TMZ treatment. Taken together, our study indicates that the CTGF knockdown and TMZ treatment effectively prevents cell invasion, migration, angiogenesis, and growth of glioblastomas in vivo. Therefore, CTGF knockdown during TMZ treatment offers a novel and potential therapeutic strategy for controlling the growth of glioblastomas. This investigation was supported in part by the R01 grants (CA-91460 and NS-57811) from the National Institutes of Health (Bethesda, MD).

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 681.
Abstract 683: Evaluating the therapeutic potential of a peptide derivative of cancer-associated PCNA in DNA repair deficient breast cancers
Authors: Phipps Elizabeth A. , Dobrolecki Lacey E. , Hickey Robert J. , Sledge George W. , Herbert Brittney-Shea , Malkas Linda H.
Published: February 2011

Our laboratory was the first to isolate and purify an intact mammalian cell multiprotein DNA replication complex, termed the synthesome, from a variety of human cell types that was both stable and fully functional. Subsequent work revealed that the synthesome of malignant breast epithelial cells exhibits a more error-prone synthesis process than that of nonmalignant breast epithelial cells. We recently correlated the low replication fidelity of malignant cells was due at least in part to a post-translational modification of a specific protein component of the synthesome: proliferating cell nuclear antigen, or PCNA. Our laboratory developed an antibody (cancer-associated or caPCNAab) specific to the cancer-associated PCNA isoform (cancer-associated or caPCNA), providing hope of an effective diagnostic marker of malignancy. Additionally, peptides were derived from the caPCNA sequence (caPeptides) and these peptides were shown to augment the action of doxorubicin (DOX) in leukemia cells while leaving nonmalignant cells intact. caPeptides may possess anti-cancer therapeutic potential through blocking the association of caPCNA with its binding partners involved in DNA replication and repair. The objective of this study was to determine the mechanism by which caPeptides directed against caPCNA induced cancer cell cytotoxicity. Breast cancer cells from a patient harboring a hereditary mutation in BRCA1, a major component of DNA double strand break repair (HCC1937 cells), were used in this study. As a genetically matched control, the wild-type breast cancer 1 early onset gene (BRCA1) was transduced into HCC1937 cells (referred to as HCC1937 + wild-type BRCA1). MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assays were conducted with caPeptides at a variety of timepoints both alone and in combination with DNA damaging agent cisplatin in HCC1937 and HCC1937 + wild-type BRCA1 cells. The data suggests cells lacking wild-type BRCA1 show increased sensitivity to caPeptides in comparison to isogenic controls transduced with wild-type BRCA1. We show that caPeptides augment the action of DNA damaging agent cisplatin. Furthermore, caPeptides containing amino acid substitutions in specific sites within the peptide show enhanced cytotoxicity over wild-type caPeptides. This work may provide information useful in a clinical setting as caPeptides could benefit a particular subset of breast cancer patients. Additionally, caPeptide studies may shed light on the deregulated replication and repair processes common in cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 683.
Abstract 686: Potentiation of the cytocidal effect of streptozotocin on carcinoid neuroendocrine cells by a pharmacological inhibitor of poly(ADP-ribose) polymerase
Authors: Richard Véronique , Shah Girish M.
Published: February 2011

Carcinoid neuroendocrine tumors (CNETs) are resistant to chemotherapy and the higher level doses of single or combination chemotherapy that are effective against these cancers also cause significant side-effects to the surrounding healthy tissues; therefore it is necessary to potentiate chemotherapy of these cancers. In the advanced clinical trials for different cancers, the pharmacological inhibitors of the DNA damage-responsive nuclear enzyme poly(ADP-ribose) polymerase-1 (PARP-1 or PARP) have shown a significant promise either as monotherapy or as potentiators of other chemotherapeutic agents. Here, we examined whether PARP-inhibitor PJ-34 can potentiate the therapy of CNETs by streptozotocin (STZ), because STZ causes DNA damage that is repaired by base excision repair pathway in which PARP-1 is known to play a significant role. We used the CNDT2.5 as model NET cells to show that treatment of cells with 1 μM PJ-34 could effectively block the activation of PARP in response to 0.5 and 1 mg/ml STZ. We also show that lower dose of STZ (0.5 mg/ml), which had no impact on the viability or the clonogenicity of cells becomes cytocidal by co-treatment with PJ-34. The potentiation effect by PJ-34 was also seen at higher (1 mg/ml) doses of STZ. We are currently examining the mechanism of potentiation of STZ in NETs by PARP-inhibitors. In conclusion, our results demonstrate that PARP-inhibitor can significantly potentiate therapeutic efficacy of STZ against NETs, and thus add back the chemotherapy option against these cancers.

Acknowledgment: We are thankful to the Canadian Institutes of Health Research for the Banting & Best Graduate Scholarship to VR and the research grant support (Grant # MOP-89964) to GMS.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 686.
Abstract 692: Antitumor activity of novel structure-based chemotherapeutic agent for the intervention of colorectal cancer
Authors: Jaiswal Aruna S. , Banerjee Sanjeev , Panda Harekrushna , Bulkin Charles D. , Izumi Tadahide , Sarkar Fazlul H. , Ostrov David A. , Narayan Satya
Published: February 2011

Colorectal cancer is the second most common cause of cancer-related death in both men and women in the Western hemisphere. The success of treatment of colon cancer patients depends on matching the most effective therapeutic regimen with the characteristics of the individual patient. The primary challenge in achieving this goal is the heterogeneity of the disease. In the past 10 years, the overall survival of colon cancer patients has significantly improved with adjuvant trials of drugs; the recurrence rate over 5 years is still high. Thus, there is clearly an urgent need for the development of new chemotherapeutic drugs and strategies. The recent emerging concept to sensitize cancer cells to DNA-alkylating drugs is by inhibiting various proteins in the base excision repair (BER) pathway. We used structure-based molecular docking of DNA polymerase β (Pol-β) and identified a potent small molecular weight inhibitor (SMI), NSC-666715. We determined the specificity of this SMI for Pol-β by using in vitro activities of APE1, Fen1, DNA ligase I, and Pol-β-directed single nucleotide (SN)- and long-patch (LP)-BER. The binding specificity of NSC-666715 with Pol-β was also determined by using fluorescence anisotropy. The effect of NSC-666715 on the cytotoxicity of the DNA-alkylating drug, Temozolomide (TMZ), to colon cancer cells was determined by in vitro clonogenic and in vivo xenograft assays. The reduction in tumor growth was higher in the combination treatment relative to untreated or monotherapy treatment. NSC-666715 showed a high specificity for blocking Pol-β activity. It blocked Pol-β-directed SN- and LP-BER without affecting the activity of APE1, Fen1 and DNA ligase I. Fluorescence anisotropy data suggested that NSC-666715 directly and specifically interacts with Pol-β and interferes with binding to damaged DNA. NSC-666715 drastically induces the sensitivity of TMZ to colon cancer cells both in vitro and in vivo assays. The results further suggest that the disruption of BER by NSC-666715 negates its contribution to drug-resistance and bypasses other resistance factors, such as mismatch repair defects. Our findings provide the “proof-of-concept” for the development of highly specific and thus safer structure-based inhibitors for the prevention of tumor progression and/or treatment of colorectal cancer. Funded by NCI-NIH grants CA-097031 and CA-100247 to S.N.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 692.
Abstract 693: Histone deacetylase inhibitors attenuate epirubicin induced p53 expression
Authors: Thurn Kenneth T. , Thomas Scott , Miller Ashleigh , Munster Pamela
Published: February 2011

Preclinical and clinical studies have demonstrated that Histone Deacetylase Inhibitors (HDACIs) can act synergistically with topoisomerase II inhibitors (e.g. doxorubicin, epirubicin) to increase the induction of apoptosis in breast cancer cells. We have previously shown that HDACIs (i.e. valproic acid and suberoylanilide hydroxamic acid, SAHA/vorinostat) caused a reduction in the expression of specific chromatin maintenance genes including HP1-, DNMT1 and SMC1-5 with subsequent chromatin decondensation. Chromatin decondensation facilitates access for DNA damaging agents such, as topoisomerase inhibitors, to their DNA substrate resulting in an increase in the topoisomerase induced DNA strand breaks and cell death. Using siRNA mediated depletion of HDAC2, similar synergistic effects were observed compared to valproic acid or SAHA. This was not observed with depletion of HDAC1 or HDAC6. This suggested the specific role of HDAC2 in mediating the synergy in increased DNA damage and apoptosis with epirubicin.

However, efficient DNA strand break repair may be an alternate mechanism of resistance. To determine whether the HDAC inhibitors play a role in the inhibition of repair enzymes in addition to the increased induction of epirubicin induced strand breaks, we sought to determine the effects of HDACIs on epirubicin mediated DNA damage repair. Previous studies have shown that various HDACis down-regulate DNA repair associated genes including BRCA1, Ku70, and Rad51. Micro-array analysis performed by our group of cultured breast cancer cells treated with valproic acid or siRNA targeting HDAC2 revealed the down-regulation of the dsDNA repair genes Rad51b and Rad51c. These Rad51 paralogs have distinct functions in homologous recombination. Rad51b has been shown to associate with, and phosphorylate p53. Rad51c is able to exchange DNA strands vital for homologous recombination, and has been suggested to recruit Rad51 to sites of DNA damage. It is also suggested to be a Trp53-dependent tumor suppressor. We show that treatment of breast cancer cells with vorinostat caused a decrease in p53 protein expression. Furthermore, pre-treatment of these cells with valproic acid or SAHA significantly inhibited the induction of p53 in the presence of epirubicin. At the time points tested (0-120 minutes), this corresponded to an initial decrease in DNA repair kinetics as demonstrated by γ-H2AX expression. We hypothesize therefore that the treatment of breast cancer cells with HDACis in the presence of epirubicin results in a decreased expression of the homologous recombination Rad51 paralog proteins and downstream p53.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 693.
Abstract 696: Anti-NRP1 tumor uptake is impacted by normal tissue sinks in a tumor bearing mouse model
Authors: Bumbaca Daniela , Boswell Charles A. , Xiang Hong , Port Ruediger , Schweiger Michelle , Stainton Shannon , Bagri Anil , Theil Frank-Peter , Fielder Paul J. , Khawli Leslie A. , Shen Ben-Quan
Published: February 2011

Neuropilin-1 (NRP1) is a VEGF165 and semaphorin receptor widely expressed by vascular endothelial cells in normal tissues and on some tumor cells. Blocking NRP1 with a human IgG1 against the b domain of NRP1 has been shown to inhibit tumor growth in several preclinical models. However, due to the wide expression of NRP1 in normal tissues, the uptake of anti-NRP1 by tumor cells may be impacted by the uptake in normal tissues. The objective of this study was to determine the normal tissue uptake of anti-NRP1 and assess its impact on tumor tissue uptake in a tumor bearing mouse model. Nude mice bearing tumors with either low, HM7, or high, SKMES, NRP1 expression were dosed with [111In]-DOTA labeled anti-NRP1 with or without excess amounts of unlabeled anti-NRP1 via single IV injection. Following dosing, blood, tumor, and several normal tissues were collected at pre-selected time points for the determination of radioactivity levels using a gamma counter. In the absence of unlabeled anti-NRP1, 3% Injected Dose (ID)/ml of radioactivity was observed in the blood at 15 minutes post-dose and decreased quickly. There was minimal radioactivity seen in tumor tissue. In contrast, the majority of radioactivity was detected in the lungs and liver. With increasing amount of unlabeled anti-NRP1, the radioactivity in the lungs/liver dropped, while increasing in blood and tumor tissue. At ∼10 mg/kg unlabeled anti-NRP1, there was no additional decrease of radioactivity in these normal tissues whereas the tumor radioactivity appeared to reach its peak. Taken together, these data suggest that the lungs/liver are tissue sinks that sequester a majority of the anti-NRP1 when given at lower doses, thus hampering tumor uptake. It is important to optimize the dose and dose regimen in order to overcome this tissue sink binding and allow for a sufficient amount of antibody to be available for tumor uptake.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 696.
Abstract 697: Inhibition of Cancer Progression by Upregulation of the Opioid Growth Factor (OGF) - OGF Receptor (OGFr) Axis Using Imquimod or Low Dose Naltrexone
Authors: McLaughlin Patricia J. , Stucki Jaimon , Zagon Ian S.
Published: February 2011

Opioid growth factor (OGF, [Met5] enkephalin), is a tonically active, negative growth factor, and interacts with a nuclear-associated receptor, OGFr. The OGF-OGFr axis mediates cell proliferation by targeting cyclin-dependent kinase inhibitory pathways. Overexpression of OGFr by pharmacological modulation of several human cancer cell lines in tissue culture has been shown to result in decreased cell replication in response to either elevated endogenous OGF or exogenous administration of this inhibitory peptide. The present study examines tumor progression in nude mice whereby established tumors of human squamous cell carcinoma of the head and neck are treated by 2 methods of upregulation of the OGF-OGFr axis: imiquimod or low dosages of the opioid antagonist naltrexone (LDN; 0.1 mg/kg naltrexone). Tumor volume was monitored 3 times weekly. Some tumors were assessed for immunohistochemical levels of OGF and OGFr. Imiquimod, a promoter of OGFr gene activity, was administered topically either 1X or 3X each week. LDN was given systemically 1X or 3X per week, or daily. At the time of death, tumor volumes for the 1X and 3X imiquimod treated mice were 18 and 47%, respectively, smaller than controls, and tumor weights were decreased 36 and 43%, respectively relative to controls. All LDN treatment regimens effectively reduced tumor volume relative to mice receiving vehicle, with a regimen-related decrease in tumor growth noted throughout the study. At termination on day 35, mean tumor volumes of mice exposed to LDN were reduced from vehicle-treated controls by 35%, 61% and 84% when given 1X/week (p<0.05), 3X/week (p<0.001) and daily (p<0.001), respectively. Tumor masses at the time of termination were 42%, 60%, and 89% smaller in the 1X/week, 3X/week, and daily, respectively, LDN treatment groups relative to controls. Semi-quantitative immunohistochemical analysis of OGF levels in tumor tissue revealed marked elevations of peptide following either imiquimod or LDN therapy. OGF tissue levels were elevated 70% and 95% in tissue from mice treated with 3X/week imiquimod or 3X/week LDN, respectively, relative to mice receiving vehicle. In summary, modulation of the OGF-OGFr axis provides new and novel therapeutic strategies for the treatment of human cancers.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 697.
Abstract 701: Elevated insulin-like growth factor 1 receptor provides an essential survival signal via Bcl-xL in rhabdomyosarcoma cells as an alternative to Bcl-2 overexpression
Authors: Mayeenuddin Linnia H. , Yu Yunkai , Kang Zhigang , Helman Lee J. , Cao Liang
Published: February 2011

Insulin-like growth factors and their receptor IGF1R play important roles in growth, development, stress response, aging, and cancer. Many antibodies targeting IGF1R are in varies stages of clinical development. It is important to understand the mechanism of the antibody-mediated cancer cell growth inhibition and death, and to identify biomarkers predictive of responses that can be implemented in clinical investigations. We previously reported that IGF1R mediated cell growth suppression of rhabdomyosarcoma (RMS) in vitro and in vivo, in which the anti-proliferative activity was correlated with IGF1R levels and AKT signaling (Cao, et al., Cancer Research 2008, 68:8039). Here, we show that in some RMS cells, treatment with a humanized IGF1R antibody results in a rapid onset of mitochondrial-dependent apoptosis, including mitochondrial depolarization, cytochrome C release, and the activation of specific caspases. The antibody-induced cell death is mediated in part via AKT-dependent pathway, and both the inhibition of AKT signaling and induction of cell death by anti-IGF1R can be rescued by the introduction of a constitutively active AKT. Further analysis reveals IGF1R antibody-induced hypophosphorylation (activation) of BAD and the activation of downstream BAX. Bcl-xL was previously shown to be a key target of activated BAD. Our data indicate that cells with the transgenic expression of Bcl-xL are completely resistant to antibody-induced cell death, while still susceptible to antibody-mediated growth inhibition. Moreover, siRNA against Bcl-xL enhances anti-IGF1R-induced apoptosis. Interestingly, the examination of RMS cell lines and tumors reveals an inverse correlation between elevated IGF1R and another pro-survival gene Bcl-2 (P=0.033). Thus, our results indicate that RMS pathogenesis may involve two strategies for cell survival: 1) increasing IGF1R expression that enhances Bcl-xL activity; or 2) up-regulating Bcl-2 expression. Our study suggests a particular subgroup of the cancer with elevated IGF1R and low Bcl-2 maybe more susceptible to anti-IGF1R treatment.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 701.