bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 59: PDK1 in melanoma development and metastasis
Authors: Scortegagna Marzia , Ruller Chelsea , Feng Yongmei , Lazova Rossitza , Kluger Harriet , Li Jian-Liang , De Surya K. , Rickert Robert , Pellecchia Maurizio , Bosenberg Marcus , Ronai Ze'ev
Published: May 2015

Phosphoinositide-dependent kinase-1 (PDK-1) is a serine/threonine protein kinase that phosphorylates members of the conserved AGC kinase superfamily, including AKT and PKC, and is implicated in important cellular processes including survival, metabolism and tumorigenesis. In large cohorts of nevi and melanoma samples, PDK1 expression was significantly higher in primary melanoma, compared with nevi, and was further increased in metastatic melanoma. Selective inactivation of Pdk1 in the melanocytes of BrafV600E::Pten-/- or BrafV600E::Cdkn2a-/-::Pten-/- mice delayed the development of pigmented lesions and melanoma induced by systemic or local administration of 4-HT. Melanoma invasion and metastasis were significantly reduced or completely prevented by Pdk1 deletion. Administration of the PDK1 inhibitor GSK2334470 (PDKi) effectively delayed melanomagenesis and metastasis in BrafV600E::Pten-/- mice. Pdk1-/- melanomas exhibit a marked decrease in the activity of AKT, p70S6K and PKC. Notably, genetic inactivation or pharmacological inhibition of PDK1 were even more effective in attenuating melanoma development and metastasis of human tumors with Pten WT as in genetic model of BrafV600E::Cdkn2a-/-::Pten+/+ mice, representing an additional fraction of human tumors harboring such mutation. Gene expression analyses identified Pdk1-dependent changes in FOXO3a-regulated genes and inhibition of FOXO3a restored proliferation and colony formation of Pdk1-/- melanoma cells. PDK1 effect on FOXO3a and melanoma development and progression appears to be AKT independent. Our studies provide direct genetic evidence for the importance of PDK1, in part through FOXO3a-dependent pathway, in melanoma development and progression.

Citation Format: Marzia Scortegagna, Chelsea Ruller, Yongmei Feng, Rossitza Lazova, Harriet Kluger, Jian-Liang Li, Surya K. De, Robert Rickert, Maurizio Pellecchia, Marcus Bosenberg, Ze'ev Ronai. PDK1 in melanoma development and metastasis. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 59. doi:10.1158/1538-7445.AM2014-59
Abstract 613: Combining PI3K and 5alpha-reductase inhibitors improves the treatment response in a mouse model of PTEN-deficient prostate cancer
Authors: Kura Yurie , Velasco Marco A. De , Ando Naomi , Fukushima Emiko , Yamamoto Yutaka , Hatanaka Yuji , Shimizu Nobutaka , Nozawa Masahiro , Yoshimura Kazuhiro , Yoshikawa Kazuhiro , Nishio Kazuto , Uemura Hirotsugu
Published: May 2015

Testosterone is converted to dihydrotestosterone (DHT) by the enzymes 5alpha-reductase types 1 and 2. Inhibition of both 5alpha-reducatase enzymes with dutasteride was effective in reducing the incidence of prostate cancer in high risk patients. This suggests that suppression of DHT with dutasteride may have a potential therapeutic effect on prostate cancer. In this study, we examined the antitumor effects of dutasteride in vivo using a preclinical model of prostate cancer that is based on the PSA-Cre mediated inactivation of PTEN. Our findings show that monotherapy with dutasteride significantly inhibited prostate tumor growth by approximately 25%. Furthermore, the combination of dutasteride with the mTOR inhibitor, everolimus, improved tumor growth suppression compared to monotherapy with either drug. Treatment with dutasteride had a minimal effect on tumor cell proliferation; however, it significantly increased the apoptotic rate in tumors. Interestingly, treatment with dutasteride resulted in increased activation of PI3K/mTOR signaling and a reduction of STAT3 pathway activation. Moreover, increased activation of MAPK, which occurs after treatment with mTOR inhibitors, was effectively reduced with the co-administration of dutasteride. The findings from our study suggest that inhibitors of 5alpha-reductase types 1 and 2 enzymes may have a potential therapeutic value in PTEN deficient prostate cancers, in particular when co-administered with PI3K inhibitors.

Citation Format: Yurie Kura, Marco A. De Velasco, Naomi Ando, Emiko Fukushima, Yutaka Yamamoto, Yuji Hatanaka, Nobutaka Shimizu, Masahiro Nozawa, Kazuhiro Yoshimura, Kazuhiro Yoshikawa, Kazuto Nishio, Hirotsugu Uemura. Combining PI3K and 5alpha-reductase inhibitors improves the treatment response in a mouse model of PTEN-deficient prostate cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 613. doi:10.1158/1538-7445.AM2014-613
Abstract 616: Transcription-regulating kinases CDK8 and CDK19 as novel therapeutic targets for advanced prostate cancer
Authors: Chen Mengqian , Kaza Vimala , Liang Jiaxin , McDermott Martina , Roninson Igor
Published: May 2015

Most of castration-refractory prostate cancers (CRPC) continue to express androgen receptor (AR) that maintains its activity at low levels or even in the absence of androgen, driving tumor cell proliferation. Androgen-depleting drugs and androgen antagonists have little or no effect against AR+ cancers that are fully androgen-independent (AI). On the other hand, experimental compounds that cause AR degradation prevent AR-mediated repression of tumor-promoting genes, a tumor-suppressive effect of AR. We report a novel druggable target for CRPC, which mediates the induction but not the repression of gene expression by AR and inhibits the growth of AI prostate cancer (PCa) cells. This target is two closely related transcription-regulating kinases CDK8/CDK19 that unlike some other members of the CDK family are not required for cell cycle progression but play critical roles in several transcriptional signaling pathways. CDK8 and CDK19 are highly expressed in AR+ PCa cells and elevated under the conditions of androgen deprivation; CDK19 is overexpressed in AR+ PCa cell lines and in metastatic clinical PCa. A novel selective CDK8/19 kinase inhibitor (Senexin B) blocks ligand-induced AR-mediated transcriptional activation in androgen-dependent (AD) PCa cells but does not affect the AR-mediated gene repression. The CDK8/19 inhibitor also inhibits ligand-independent AR activation and AI cell growth in AI-PCa cells resistant to the anti-androgenic drug enzalutamide, both in vitro and in a mouse xenograft model. We have also discovered that CDK8/19 inhibition decreases NFκB mediated transcriptional activation of a group of tumor-promoting chemokines, such as CXCL1, CXCL2 and IL8, suggesting that CDK8/19 may promote prostate cancer chemoresistance and metastasis through the NFκB pathway. The above results suggest that CDK8/19 is an especially attractive therapeutic target for advanced PCa, which controls both AR-driven cell proliferation and other oncogenic pathways.

Citation Format: Mengqian Chen, Vimala Kaza, Jiaxin Liang, Martina McDermott, Igor Roninson. Transcription-regulating kinases CDK8 and CDK19 as novel therapeutic targets for advanced prostate cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 616. doi:10.1158/1538-7445.AM2014-616
Abstract 627: Immunogenic cell death as novel immune response mechanism to EGFR-targeted therapy in CRC
Authors: Penna Giuseppe , Pozzi Chiara , Magni Elena , Curigliano Giuseppe , Zampino Maria Giulia , Rescigno Maria
Published: May 2015

Colorectal cancer (CRC) is the third most common cancer worldwide. The major improvement in the median overall survival of mCRC patients was mainly due to the introduction of new active chemotherapeutic and biomolecular targeting agents. Most chemotherapeutic agents and radiotherapy kill tumor cells through an apoptotic pathway, which is considered as a non-immunogenic. However, anthracyclines, oxaliplatin or ionizing irradiation are able to induce a potent immune response in vivo, when injected in immunocompetent mice. Dying cells are readily taken up by dendritic cells (DC) that then cross-present their associated antigens for effective T cell activation.

Herein we propose that the treatment with an anti-EGFR antibody (Cetuximab) in combination with chemotherapy FOLFIRI (5-fluorouracile, leucovorin, irinotecan) induces a immunogenic cell death (ICD) modality, which contributes to its clinical efficacy in patients with mCRC expressing EGFR and the wild type (wt) form of the viral oncogene v-Ki ras2.

Our results, obtained on CRC cell lines provide evidence, for the first time, that treatment with Cetuximab alone or in combination with a chemotherapeutic regimen (FOLFIRI) directs dying cells towards an ICD process characterized by phosphorylation of eIF2α and, most important, increased phagocytosis by DCs. However, not all cell lines wt for KRAS (expressing EGFR) displayed the same response. As expected, Cetuximab blocked the pathways leading to AKT and ERK phosphorylation. However, we also observed that Cetuximab, and to a lesser extent the F(ab)2 fragments of Cetuximab, was inducing some phosphorylation of the EGFR in KRAS wt responding cell line, which was lower than that induced by EGF . Hence Cetuximab is uncoupling the activation of the signaling pathway induced via EGFR from EGFR phosphorylation, suggesting an alternative activation of the receptor. This allowed us to postulate that Cetuximab-induced phagocytosis may be the effect of an alternative activation of the EGFR. Indeed, we could not detect any phagocytosis in cell lines where the phosphorylation of the receptor did not occur in response to Cetuximab.

These results identify ICD as novel immune response mechanism to EGFR targeted therapy alone or in combination with chemotherapy and possibly genetic alterations of the EGFR-signalling pathway, could interfere with the induction of ICD.

Citation Format: Giuseppe Penna, Chiara Pozzi, Elena Magni, Giuseppe Curigliano, Maria Giulia Zampino, Maria Rescigno. Immunogenic cell death as novel immune response mechanism to EGFR-targeted therapy in CRC. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 627. doi:10.1158/1538-7445.AM2014-627
Abstract 630: Induction of anti-melanoma immunity after intralesional ablative therapy
Authors: Liu Hao , Kodumudi Krithika , Weber Amy , Sarnaik Amod A. , Pilon-Thomas Shari
Published: May 2015

Immunotherapeutic strategies incorporating intralesional (IL) ablative therapy to elicit a tumor specific immune response are under investigation as a non-surgical option to induce tumor regression of cutaneous neoplasms. Rose Bengal (RB) is a water-soluble xanthene dye that was originally used as an intravenous liver diagnostic and is in use by ophthalmologists to stain damaged cells in the eye. In murine models of breast cancer and melanoma, we have shown that IL injection of PV-10 (10% RB in saline solution) leads to ablation of injected tumors and regression of non-injected bystander tumors. In these models, increased anti-tumor T cell responses were measured, supporting the induction of systemic anti-tumor immunity after tumor ablation with PV-10. In our ongoing phase I clinical trial exploring melanoma regression in patients, IL PV-10 has led to a significant decrease of melA positive melanoma cells in the biopsies of both PV10-injected and non-injected lesions. This regression correlated with increased circulating CD3+T cells (p=0.03) in peripheral blood mononuclear cells (PBMC). T cells purified from PBMC from a melanoma patient produced increased IFN-gamma in response to autologous tumor after treatment with PV-10. Ex vivo models implemented to investigate this phenomenon indicate that the cytotoxicity induced by PV-10 is not apoptosis-dependent as evidenced by Annexin staining of melanoma cells following PV-10 treatment. PV-10 directly induced necrosis of melanoma cells at 50 uM, but was not toxic to healthy fibroblasts at the same dose. Further preclinical translational testing has shown that treatment of murine B16 cells with PV-10 leads to release of HMGB1, a soluble Damage Associated Molecule Pattern (DAMP) that is important for activation of dendritic cells (DCs). In the murine B16 melanoma model, there is a significant increase in the number of DCs infiltrating the tumor-draining lymph nodes after IL injection of PV-10. These findings suggest that PV-10 treatment leads to the release of DC activating factors and DC recruitment. Further studies to determine the role of PV-10 on T cell activation are ongoing. In sum, these clinical and preclinical results increase our understanding of the cytotoxic and immunological mechanisms that may play a role in systemic immunity induced by PV-10 tumor ablation.

Citation Format: Hao Liu, Krithika Kodumudi, Amy Weber, Amod A. Sarnaik, Shari Pilon-Thomas. Induction of anti-melanoma immunity after intralesional ablative therapy. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 630. doi:10.1158/1538-7445.AM2014-630
Abstract 641: Targeted inhibition of BRAF signaling in melanoma enhances the activity of a multivalent immunotherapeutic vaccine
Authors: Suriano Robert , Tuli Neha , Geliebter Jan , Tiwari Raj K. , Wallack Marc K.
Published: May 2015

Development of a successful immunotherapeutic vaccine targeting melanoma requires the use of multiple melanoma associated antigens (MAA). Expression of several MAAs is regulated by the micropthalmia transcription factor (MITF), which in turn is regulated by members of the MAPK pathway, such as ERK. It is also known that 70% of melanomas express the genetic lesion BRAFV600E. Based on this knowledge, we hypothesize that the use of a specific BRAFV600E inhibitor would enhance MITF mediated MAA expression and thus serve as a viable target for immunotherapy. We have developed a vaccinia virus based melanoma vaccine using patient derived primary melanoma cells that are rich in MAAs such as gp100, tyrosinase, and TRP-1. Our results indicated that our cells were either homozygous or heterozygous for the mutated BRAF gene. In addition, our cells were responsive to treatment by PLX-4032, an FDA approved treatment for melanoma, as evidenced by inhibition of phosphorylation of MEK and ERK, two key players in the MAPK pathway. Phenotypically, the migratory and invasive properties of our cell lines were also altered by PLX-4032. Most interestingly was the observation that upon treatment with PLX-4032, the expression of various MAAs was altered. Specifically, the expression of gp100, MART-1, and tyrosinase increased both on the RNA level as well as on the protein level. Our results warrant further investigation into a combinatorial therapeutic approach to treat melanoma by which increasing the expression of various immunologically relevant MAAs, upon treatment with PLX-4032, followed by vaccination may lead to a more robust and targeted anti-melanoma immune response.

Citation Format: Robert Suriano, Neha Tuli, Jan Geliebter, Raj K. Tiwari, Marc K. Wallack. Targeted inhibition of BRAF signaling in melanoma enhances the activity of a multivalent immunotherapeutic vaccine. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 641. doi:10.1158/1538-7445.AM2014-641
Abstract 673: A Monoclonal Antibody, MORAb-075, Targeting Claudin-3 & Claudin-4 Inhibits human ovarian, pancreatic and prostate cancer progression in mice
Authors: Lin Jian-Min , soto allis , maddage Christopher , Albone earl , zhou yuhong , grasso luigi
Published: May 2015

Claudin (CLDN) is a family of integral membrane proteins. They are crucial for tight junction formation and functions. Studies have found CLDN3 and CLDN4 proteins are overexpressed in majority of primary ovarian, prostate, and pancreatic cancers.

We have generated a rat-human chimeric monoclonal antibody, MORAb-075, which binds to CLDN3 and CLDN4 with high affinity in vitro. In order to evaluate its therapeutic potential in vivo, we have established a number of human cancer models where the tumors are grown orthotopically in nude mice. These tumors were confirmed positive for human CLDN3 or/and CLDN4 expressions via immunohistochemistry staining. The models include a CLDN3/4-expressing prostate cancer PC-3; a CLDN4-expressing pancreatic cancer, BxPc-3; a CLDN3-expressing ovarian cancer, IGROV-1. In these orthotopic tumor models, MORAb-075 demonstrated significant antitumor activities. When the mAb was administered intravenously at 25 mg/kg, twice weekly for 4 weeks, the tumor growth inhibition (TGI) of PC-3 tumors reached 50%; TGI of BxPC-3 tumors was nearly 60% and TGI of IGROV-1 tumors was greater than 75%. Our findings demonstrate the potency of MORAb-075, as a single agent, in inhibiting solid tumor growth in these models. These results suggest that mAb therapy targeting CLDN3 & CLDN4 is a valuable option for therapeutic application in cancer therapy that may be improved in combination with standard of care chemotherapies for CLDN3 or CLDN4 positive cancers.

Citation Format: Jian-Min Lin, allis soto, Christopher maddage, earl Albone, yuhong zhou, luigi grasso. A Monoclonal Antibody, MORAb-075, Targeting Claudin-3 & Claudin-4 Inhibits human ovarian, pancreatic and prostate cancer progression in mice. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 673. doi:10.1158/1538-7445.AM2014-673
Abstract 680: HMGB1 regulates autophagy and apoptosis to promote chemotherapy resistance in multiple myeloma
Authors: Zhang Xingding , Qi Lin , Yang Lin
Published: May 2015

Multiple myeloma is the second most frequently diagnosed hematologic malignancy in the United States after non-Hodgkin lymphoma. Unfortunately, multiple myeloma remains incurable, and virtually all patients eventually relapse with disease that is typically more resistant to therapy than in prior lines of treatment, for which the underlying molecular mechanisms remain unclear. In this study, we showed that high-mobility group box 1 (HMGB1), the best characterized damage-associated molecular pattern, in the development of chemotherapy resistance in multiple myeloma. To examine its potential role of chemotherapy resistance in myeloma, we detected protein expression of HMGB1 treated with anticancer chemotherapy agent doxorubicin and observed that HMGB1 was upregulated by different concentrations of doxorubicin in myeloma cells. Chemosensitivity assay was performed with RPMI 8226 and U266 cells with knockdown/overexpression of HMGB1 and treated with doxorubicin, and cells were detected for their viability by WST-1 assay after 72 hr. Consistently, shRNA interference-mediated knockdown of HMGB1 cells exhibited significant sensitized to doxorubicin treatment compared to non-targeting shRNA cells, and HMGB1-overexpressing cells significantly resistance to doxorubicin. Furthermore, studies in xenograft models showed that shRNA of HMGB1 also sensitized the chemosensitivity of multiple myeloma cells in vivo. At the molecular level, mechanistic investigation showed that shRNA of HMGB1 decreased autophagy, which is an intracellular self-defense mechanism to drug resistance, by inhibition of the autophagic marker microtubule-associated protein light chain 3-II and increasing of sequestosome 1 (p62) and autophagosome formation. Finally, we found that HMGB1 regulated autophagy and apoptosis by binding to p53 and Beclin1, and p53 is a negative regulator of the HMGB1/Beclin1 complex. Taken together, these data support the hypothesis that HMGB1 modulates chemotherapy resistance in myeloma through effects on the pathway of autophagy and apoptosis, as well as a target for strategies to aim at laying the foundation for further clinical studies, and overcome chemoresistance in multiple myeloma.

Note: This abstract was not presented at the meeting.

Citation Format: Xingding Zhang, Lin Qi, Lin Yang. HMGB1 regulates autophagy and apoptosis to promote chemotherapy resistance in multiple myeloma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 680. doi:10.1158/1538-7445.AM2014-680
Abstract 702: DCVex(TM): A novel integration-deficient lentivector technology that incorporates genetic and post-translational elements to target dendritic cells
Authors: Tareen Semih U. , Kelley-Clarke Brenna , Nicolai Christopher J. , Slough Megan M. , Vin Chintan D. , Hoeven Neal Van , Robbins Scott H. , Meulen Jan H. ter , Berglund Peter
Published: May 2015

With the goal of creating antigen-directed immunotherapeutics that can be safely administered directly to patients, Immune Design has developed a platform of novel integration-deficient lentiviral vectors that target and deliver antigen-encoding nucleic acids to dendritic cells (DCs) in order to promote the activation of antigen-specific effector CD8 T cells. This platform, termed DCVex(TM), utilizes a novel genetic variant of a Sindbis virus envelope glycoprotein with post-translational carbohydrate modifications in combination with Vpx, a SIVmac viral accessory protein, to achieve efficient targeting and transduction of DCs. In addition, DCVex(TM) incorporates safety features in its design that include redundant mechanisms to render DCVex(TM) integration-deficient, as well as genetic modifications that eliminate psi-gag recombination between split genome components. The characteristics that allow DCVex(TM) to specifically transduce human DCs and the advances that DCVex(TM) brings to conventional third-generation lentiviral vector design demonstrate its potential as a vaccine designed to utilize DCs for cancer immunotherapy.

Citation Format: Semih U. Tareen, Brenna Kelley-Clarke, Christopher J. Nicolai, Megan M. Slough, Chintan D. Vin, Neal Van Hoeven, Scott H. Robbins, Jan H. ter Meulen, Peter Berglund. DCVex(TM): A novel integration-deficient lentivector technology that incorporates genetic and post-translational elements to target dendritic cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 702. doi:10.1158/1538-7445.AM2014-702
Abstract 704: Comparison of Salmonella typhimurium A1-R and VNP20009 on the Lewis Lung carcinoma
Authors: Zhang Yong , Zhang Nan , Hoffman Robert M. , Zhao Ming
Published: May 2015

Salmonella typhimurium A1-R (A1-R) is auxotrophic for arg and leu, which attenuates growth in normal tissue but allows high tumor virulence. A1-R is effective against metastatic human prostate, breast, and pancreatic cancer as well as osteosarcoma, fibrosarcoma, and glioma in clinically-relevant mouse models. VNP20009 is also a genetically-modified strain of Salmonella typhimurium that has been tested in a Phase I clinical trial, but has more attenuation mutations than A1-R. Lewis-lung mouse lung cancer-bearing mice were treated with A1-R and VNP20009 in nude mice. A1-R and VNP20009 were both eliminated from the liver and spleen approximately 3-5 days after administration via tail vein, however A1-R showed higher tumor targeting and increased mouse survival than VNP20009 with less body weight loss. The mice tolerated A1-R at least 2-fold higher than NVP20009 when bacteria were administered iv. The results of the present study suggest that A1-R has greater clinical potential than VNP20009.

Citation Format: Yong Zhang, Nan Zhang, Robert M. Hoffman, Ming Zhao. Comparison of Salmonella typhimurium A1-R and VNP20009 on the Lewis Lung carcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 704. doi:10.1158/1538-7445.AM2014-704
Abstract 715: Vivo-Morpholino antisense oligomers decrease tumor growth in mice by altering mVEGF mRNA splicing to knock down mVEGF expression
Authors: Jiang Shan
Published: May 2015

Vivo-Morpholino antisense oligomers can knock down gene expression by altering splicing of the pre-mRNA and generating a translational frameshift. VEGF overexpression is crucial for tumor growth and anti-VEGF therapies are promising in the treatment of certain cancers. Decreasing VEGF gene expression by exon deletion using Vivo-Morpholinos could suppress tumor growth. We examine the efficiency of mVEGF exon 2 deletion and tumor growth inhibition in mice injected with a Vivo-Morpholino oligomer (VMO-e2i2), which is designed to specifically delete the mVEGF exon 2 and cause downstream frameshifting in mVEGF protein. Mice bearing s.c. Lewis lung carcinoma tumors are I.V. injected with the VMO-e2i2 oligomers at 15mg/kg twice weekly for 2 weeks. VEGF exon 2 deletions are detected by RT-PCR in various tissues tested: approximately 100% exon 2 deletion in kidney, 80% in liver, 70% in skeletal muscles and 40% in s.c. tumors. The tumor volumes decrease by 6 fold over 16 days in mice injected with the VMO-e2i2 oligomers. Exon deletions are detected in various tissues including liver, small intestine, colon, skeletal and cardiac muscles, lung, stomach and kidney. The exon deletion mediated by Vivo-Morpholinos is target specific, dose-dependent and is detectable for more than 15 weeks after a single IV injection in mice. No significant toxicity is observed in mice injected with the effective dosage. Vivo-Morpholinos therefore represent a useful tool for cancer target identification and validation in rodent models and a possible therapeutic treatment.

Citation Format: Shan Jiang. Vivo-Morpholino antisense oligomers decrease tumor growth in mice by altering mVEGF mRNA splicing to knock down mVEGF expression. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 715. doi:10.1158/1538-7445.AM2014-715
Abstract 736: DJ4, a novel ROCK and MRCK inhibitor, potently inhibits migration and invasion of cancer cells
Authors: Kale Vijaykumar P. , Desai Dhimant , Dick Taryn , Choe Katherine , Colledge Ashley , Amin Shantu , Hengst Jeremy , Yun Jong
Published: May 2015

Rho-associated coiled-coil containing protein kinase (Rho kinase/ROCK) and myotonic dystrophy kinase-related Cdc42-binding kinase (MRCK) are involved in the aggressive migration of cancer cells. These are the key proteins involved in the formation of stress fibers that confer contractility to cancer cells during migration and invasion. ROCK and MRCK are primarily involved in amoeboid and mesenchymal type of migration of cancer cells respectively. Earlier studies reported that combined inhibition/silencing of ROCK and MRCK has more pronounced effect than inhibition of the individual kinases suggesting a more effective therapeutic strategy to combat metastatic cancers. With this hypothesis in mind, we developed a series of novel multikinase inhibitors and identified DJ4 as a potent ATP competitive inhibitor of ROCK1, ROCK2 and MRCKβ. DJ4 effectively inhibits the kinase activity of ROCK (ROCK1 and ROCK2) and MRCK in a cell-free system and multiple human non-small cell lung cancer cell lines. In cellular functional assays DJ4 inhibited migration of human melanoma, lung, pancreatic and breast cancer cells in a concentration-dependent manner. In live cell imaging over 20h by time-lapse microscopy, DJ4 (2.5µM ) reduced the migration rate of lung-and breast-cancer cells by 2.7 and 5.5 times compared to control cells respectively. DJ4 (5µM) also inhibited invasion of MDA-MB-231 breast cancer cells by ∼70%. Fluorescent microscopic observation also revealed that inhibition of activity of these kinases, by DJ4, causes reduction in stress fibers which leads to inhibition of cell contractility and migration.

In summary, our studies demonstrate that DJ4 is a potent ATP competitive inhibitor of ROCK and MRCK, and effectively inhibits migration and invasion of multiple cancers in vitro. Together these results indicate that DJ4 has a strong potential to be an anti-metastatic drug against multiple cancer types.

Citation Format: Vijaykumar P. Kale, Dhimant Desai, Taryn Dick, Katherine Choe, Ashley Colledge, Shantu Amin, Jeremy Hengst, Jong Yun. DJ4, a novel ROCK and MRCK inhibitor, potently inhibits migration and invasion of cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 736. doi:10.1158/1538-7445.AM2014-736
Abstract 752: Genetic and pharmacological FAK inhibition disrupt a β5 integrin signaling axis controlling anchorage-independent ovarian carcinoma growth
Authors: Tancioni Isabelle , Uryu Sean , Sulzmaier Florian , Shah Nina , Lawson Christine , Miller Nichol L.G. , Jean Christine , Chen Xiao Lei , Ward Kristy K. , Schlaepfer David D.
Published: May 2015

Ovarian cancer spreads via cell shedding and growth within malignant ascites. Effective targeted therapies have not been developed for ovarian cancer. Ascites contains an abundance of matrix proteins, and spheroids maintain integrin receptor expression. Through databases analyses we find that elevated osteopontin (OPN), β5 integrin, and focal adhesion kinase (FAK) mRNA levels are associated with decreased overall survival of serous ovarian cancer patients treated with platinum and taxol. In ovarian tumor tissue arrays, increased FAK activation (FAK Y397 phosphorylation) correlated with elevated tumor grade in parallel with increased in β5 integin and OPN levels. FAK is a cytoplasmic tyrosine kinase that remains active in spheroids, and treatment of seven ovarian carcinoma cell lines with sub-micromolar levels of FAK inhibitor (PND-1186) identified sensitive (HEY and OVCAR8), intermediate (OVCAR3, ID8-IP, and IGROV1-IP), and resistant (SKOV3-IP and OVCAR10) cells to blockage of growth under anchorage-independent conditions. Genetic or pharmacological FAK inhibition within ID8-IP or HEY cells selectively prevents anchorage-independent growth in culture and tumor growth in mice with corresponding reductions in β5 integrin and OPN expression. β5 knockdown reduced HEY growth in soft agar, tumor growth in mice, FAK Y397 phosphorylation, and OPN expression in spheroids. Although FAK inhibitor resistant ovarian carcinoma cells (SKOV3-IP and OVCAR10) were associated with anchorage-independent Akt S473 phosphorylation, membrane-targeted and activated Akt expression in sensitive cells (HEY and OVCAR8) resulted in only a partial rescue of FAK inhibitor-associated growth block. These results support the hypothesis that OPN, αvβ5 integrins, and FAK may function as a signaling axis promoting ovarian tumor progression. Although Akt signaling pathway activation is a common event in serous ovarian cancer, our results suggest that this may not impart complete resistance to FAK inhibitor treatment.

Supported by NIH CA102310

Citation Format: Isabelle Tancioni, Sean Uryu, Florian Sulzmaier, Nina Shah, Christine Lawson, Nichol L.G. Miller, Christine Jean, Xiao Lei Chen, Kristy K. Ward, David D. Schlaepfer. Genetic and pharmacological FAK inhibition disrupt a β5 integrin signaling axis controlling anchorage-independent ovarian carcinoma growth. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 752. doi:10.1158/1538-7445.AM2014-752
Abstract 754: Selective ALK inhibitor alectinib (CH5424802/RO5424802) with potent antitumor activity in models of crizotinib resistance, including intracranial metastases
Authors: Kodama Tatsushi , Tsukaguchi Toshiyuki , Hasegawa Masami , Yoshida Miyuki , Takanashi Kenji , Kondoh Osamu , Sakamoto Hiroshi
Published: May 2015

Purpose: EML4-ALK has been implicated as a driver oncogene and a therapeutic target in non-small cell lung cancer (NSCLC). The clinical efficacy of the ALK inhibitor crizotinib has been demonstrated in ALK fusion-positive NSCLC; however, resistance to crizotinib certainly occurs through ALK secondary mutations in clinical use. In addition, brain metastases are frequent sites of initial failure in patients because crizotinib has poor penetration of the central nervous system. This study aimed to clarify the efficacy of ALK inhibitor alectinib (CH5424802/RO5424802) in models of crizotinib-resistant ALK mutant tumors and intracranial tumors.

Experimental Design: The antitumor activity of alectinib was evaluated in subcutaneous xenograft tumor models of Ba/F3 cells expressing mutated EML4-ALK, and in intracranial tumor implantation models of EML4-ALK-positive NSCLC NCI-H2228.

Results: Alectinib had kinase inhibitory activity against some ALK mutations, including G1269A, and blocked tumor growth driven by a mutant of EML4-ALK in vitro and in vivo. In intracranial tumor implantation mouse models, alectinib resulted in regression of NCI-H2228 tumor in brain and provided a survival benefit. Alectinib is not a substrate for P-glycoprotein efflux transporter in vitro and showed a high brain-to-plasma ratio in rat. Additionally, alectinib led to tumor size reduction in the subcutaneous NCI-H2228 xenograft tumor that had failed to regress fully during treatment with crizotinib.

Conclusions: Alectinib was effective against most EML4-ALK mutations and showed potent efficacy against intracranial ALK-positive tumor. Thus, alectinib might provide therapeutic opportunities for crizotinib-treated patients with ALK secondary mutations and brain metastases.

Citation Format: Tatsushi Kodama, Toshiyuki Tsukaguchi, Masami Hasegawa, Miyuki Yoshida, Kenji Takanashi, Osamu Kondoh, Hiroshi Sakamoto. Selective ALK inhibitor alectinib (CH5424802/RO5424802) with potent antitumor activity in models of crizotinib resistance, including intracranial metastases. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 754. doi:10.1158/1538-7445.AM2014-754
Abstract 757: ET-D5, first-in-class synthetic selective Protein Phosphatase 1 (PP1) inhibitor with anti-proliferative and anti-vascular activity
Authors: Juhem Aurelie , Lecercle Delphine , Deverchère Julie , Gazel Arnaud , Florent Jean-Claude , Popov Andrei
Published: May 2015

In this work we describe ET-D5, a synthetic small molecule serine-threonine protein phosphatase inhibitor with proven in vivo anti-mitotic and anti-vascular properties.

The potential anti-cancer properties of serine-threonine protein phosphatase inhibitors have been identified since ancient times (extract from blister beetle mylabris was used in China for cancer treatment as early as in 1264), and several, mostly natural, inhibitors were evaluated in clinical trials. Their development was, however, discontinued due to the toxic effects of these non-selective (PP2A>>PP1) serine-threonine protein phosphatase inhibitors.

ET-D5 is a remarkably specific (PP1 versus PP2A) inhibitor with potent anti-mitotic and anti-vascular activities. In vitro, ET-D5 display high efficacy against a wide range of human cancer cells (GI50 values determined were in low nM range for certain cell lines). In addition, ET-D5 disrupts the in vitro preformed capillary tubes (endothelial cells HMEC-1). In vivo experiments confirmed these activities in numerous animal models of human cancer, such as cancer of lung, brain, thyroid, kidney and ovaries.

ET-D5 presents interesting pharmacological properties, good toxicology profile and is also active when administered per os. Contrary to many vascular-disrupting agents (VDA) which target tubulin, ET-D5 exerts its activity by selectively inhibiting in vivo protein-phosphatase I (PP1). We confirmed this activity in classical pharmacodynamics experiments, showing PP1 inhibition in xenografted tumors after oral administration of ET-D5.

Based on all these results, ET-D5 is scheduled to enter clinical trials in early 2015.

Citation Format: Aurelie Juhem, Delphine Lecercle, Julie Deverchère, Arnaud Gazel, Jean-Claude Florent, Andrei Popov. ET-D5, first-in-class synthetic selective Protein Phosphatase 1 (PP1) inhibitor with anti-proliferative and anti-vascular activity. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 757. doi:10.1158/1538-7445.AM2014-757
Abstract 76: Liver receptor homologue-1 increases incidence of DMBA-induced mammary tumors
Authors: Lazarus Kyren A. , Brown Kristy , Young Morag , Cain Jason , Jayasekara Samantha , Coulson Rhiannon , Watkins Neil , Clyne Colin , Chand Ashwini
Published: May 2015

Breast cancer is one of the most common malignancies globally. It accounts for approximately 15% of cancer-related deaths in Australian women. The orphan nuclear receptor liver receptor homolog-1 (LRH-1) promotes increased cell proliferation, motility and invasion in breast cancer cell lines. Additionally, high LRH-1 expression in human breast cancers is positively associated with estrogen receptor alpha status and aromatase activity. However, the role of LRH-1 in tumour growth is not well understood. Therefore, we aimed to generate a doxycycline (dox)-inducible mammary epithelial specific LRH-1 knock-in mouse in order to define the role of LRH-1 in mammary epithelial proliferation in vivo. In addition, the Dimethylbenz(a)anthracene (DMBA) induced mammary tumour model was utilized with the LRH-1 transgenic mice to determine the role LRH-1 plays in promoting mammary carcinogenesis. Given clear in vitro roles for LRH-1 in breast cancer cell proliferation, we hypothesise LRH-1 stimulates mouse mammary epithelial cell proliferation and promotes DMBA-mediated mammary tumorigenesis.

We demonstrate increase in proliferation markers Ki-67, PCNA and PH3 immunoreactivity in luminal epithelial cells of dox-treated animals indicating that LRH-1 plays a role in mammary cell proliferation in vivo. In DMBA treated animals, we observed a five-fold increase of dense pre-neoplastic epithelial foci (p=0.0267), and a four-fold increase in microscopic lesions in dox-treated animals (p=0.0003). We also demonstrated that LRH-1 over-expression significantly reduced breast tumour-free survival in the transgenic DMBA model (no dox n=11; dox n=12, Mantel-Cox test p=0.0375). Tumour penetrance in DMBA animals not treated with dox was 9% (out of eleven animals) versus 25.0% (out of twelve animals) in the dox treated cohort.

LRH-1 is known synergise with β-catenin to induce cyclin D1/E1 mediated cell proliferation in vitro. We analysed transcript levels of cyclin D1/E1 and show a significant increase in dox treated animals with or without DMBA. Finally, we demonstrate an increase in the oncogene β-catenin nuclear localisation in the dox treated animals with or without DMBA. Taken together, these data suggest that LRH-1 increases incidence of DMBA-induced mammary tumours. Further analyses on mechanism(s) mediated by LRH-1 are warranted to fully understand its role in mammary tumorigenesis.

Note: This abstract was not presented at the meeting.

Citation Format: Kyren A. Lazarus, Kristy Brown, Morag Young, Jason Cain, Samantha Jayasekara, Rhiannon Coulson, Neil Watkins, Colin Clyne, Ashwini Chand. Liver receptor homologue-1 increases incidence of DMBA-induced mammary tumors. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 76. doi:10.1158/1538-7445.AM2014-76
Abstract 764: A role for GLI1 in the development of multidrug resistance in rhabdomyosarcoma (RMS) cells
Authors: Yoon Joon Won , Lamm Marilyn , Leong King-Fu , Iannaccone Stephen , Iannaccone Philip , Walterhouse David
Published: May 2015

RMS is the most common sarcoma of childhood. About 30% of patients with localized tumors will recur following treatment. The outcome for patients with recurrent RMS remains poor. Therefore, development of chemotherapy resistance during RMS therapy represents an important problem and novel approaches to prevent or reverse drug resistance are essential. Activation of multidrug transporter genes, including MDR1, MRP1, LRP and TAP1 represents an important mechanism for drug resistance in RMS. However, the mechanism of expression of multidrug resistance genes in RMS is incompletely understood. Recent reports have suggested a role for the Hedgehog (HH) signaling pathway and its downstream mediator GLI1 in acquisition of a multidrug resistance phenotype in esophageal adenocarcinoma, glioblastoma, and myeloid leukemia. Since approximately 60% of embryonal RMS (ERMS) and more rarely alveolar RMS (ARMS) express HH pathway components, we hypothesized that HH pathway activation may up-regulate the expression of multidrug resistance genes in RMS. We demonstrated expression of HH pathway components, including GLI1, in ERMS (RD, Rh18 and Ruch-2) and ARMS (Rh30 and Rh41) cell lines by qRT PCR and/or Western blot. We found variable expression of MDR1, MRP1, LRP and TAP1 by qRT PCR in these cell lines. Treatment of RMS cells with GLI1 siRNA (RD, Rh18 and Rh41) or the GLI1 antagonist GANT61 (Rh18 and Rh30) caused down-regulation LRP and TAP1 expression. In addition, GLI1, MDR1, LRP and TAP1 expression was increased in vincristine resistant UKF Rhb-1 RMS cells compared with parental cells (obtained from Dr. Cinatl) by qRT PCR. Treatment of vincristine resistant UKF Rhb-1 cells with vincristine together with either GANT61 or Cpd#33 (GLI1 inhibitors), each at a dose, which did not reduce cell viability individually, significantly decreased cell viability by MTT assay. Our results suggest that GLI1 up-regulates a subset of multidrug resistance genes in RMS cells and that GLI1-inhibitors may reduce multidrug resistance.

Citation Format: Joon Won Yoon, Marilyn Lamm, King-Fu Leong, Stephen Iannaccone, Philip Iannaccone, David Walterhouse. A role for GLI1 in the development of multidrug resistance in rhabdomyosarcoma (RMS) cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 764. doi:10.1158/1538-7445.AM2014-764
Abstract 779: Osteopontin gene expression is associated with 5-fluorouracil drug resistance in colon cancer
Authors: Nakajima Go , Kuramochi Hidekazu , Yamamoto Masakazu , Hayashi Kazuhiko
Published: May 2015

Background: Resistance to chemotherapeutic drugs complicates the treatment of cancer patients. The extracellular matrix protein osteopontin (OPN) plays multiple roles in the proliferation and metastasis of cancer cells. We therefore attempted to determine whether OPN expression correlated with drug resistance. Methods: OPN expression in the HCT 116 colon cancer cell line was inhibited by an OPN-short interfering RNA (siRNA). We determined the cytotoxic effect (IC50) of 5-fluorouracil (5FU) on these cells. Patients with recurrence or colorectal cancer or harbored residual tumor cells were treated with S-1-based chemotherapy. The levels of OPN mRNA expression in the tumors were determined and compared with the patients’ responses to chemotherapy. Results: The IC50 of 5FU for HCT 116 cells transfected with the OPN-siRNA was 32.9 μM. In contrast, the IC50 values for cells transfected with a negative control siRNA, mock-transfected cells, or untreated cells were 6.58 μM, 7.08 μM, and 6.76 μM, respectively (P<0.001). The level of OPN mRNA expression in the S-1 non-responder group was significantly lower than that of the responder group (P = 0.0387). Survival analysis revealed no significant difference between the responder and non-responder groups (P = 0.8737). Conclusions: The level of OPN expression plays a role in the resistance of colorectal tumor cells to 5FU.

Citation Format: Go Nakajima, Hidekazu Kuramochi, Masakazu Yamamoto, Kazuhiko Hayashi. Osteopontin gene expression is associated with 5-fluorouracil drug resistance in colon cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 779. doi:10.1158/1538-7445.AM2014-779
Abstract 780: Metformin induces apoptotic cell death and inhibits CD133+ cancer stem cells in 5-Fu-resistant colorectal cancer cells
Authors: Kim Sunghee , Ku Ja-Lok
Published: May 2015

Colorectal cancer (CRC) is the third most common cancer and estimated cancer related death is high in worldwide. 5-Fluorouracil (5-Fu) is common used chemotherapeutic agent for colorectal cancer. Nevertheless the use frequency, the resistance to 5-Fu is still a critical problem. To improvement of therapeutic effects of 5-Fu, there are the new chemotherapeutic strategies like FOLFOX and FORFIRI, combination treatment 5-Fu with oxaliplatin or irinotecan. However, it is arguable that the side effects and efficiency of these strategies.

Metformin is widely used therapeutic agent for type II diabetics, obesity, and polycystic ovarian syndrome with limited side effects. Recently, there are some evidence that metformin has anticancer effects for some types of cancers e.g. breast, pancreases, prostate, and colon. Recently, correlation of metformin and cancer stem cell population was proved.

Here, we examined that metformin effects on parental colorectal cancer cells and their 5-Fu-resistant colorectal cancer cells. Metformin has the anticancer effects in parental cells and their 5-Fu-resistant cells in both cell lines. As treated with metformin, wound healing ability is decreased in both cell lines. Cell proliferation was 41.7% decreased in parental cells and 30.6% in 5-Fu-resistant cells. Also, expression of PARP and caspase-3 were induced by metformin dose dependent manner in both cell lines. It is suggested that metformin induces inhibition of cell proliferation and increased apoptosis. In addition, metformin treatment affects cell cycle; subG1 stage was increased to 1.7 fold and G2/M stage was increased to 1.6 fold in both cell lines. It means that metformin induces cell death and leads cell cycle arrest. However, there are no significant synergistic effect in cell proliferation, apoptosis, and cell cycle change with combination treatment of 5-Fu and metformin. CD133+ cancer stem cell population was decreased in 20 to 40 % as treated with 5-Fu and low dose of metformin to parental cells. On the other hand, CD133+ population in 5-Fu-resistant cells was affected in only low dose of metformin not 5-Fu.

In sum, metformin has anticancer effects via inhibition of cell proliferation, cell cycle changes and metformin decreases CD133+ cancer stem cell population. Further studies will be necessary to verify the mechanisms that metformin decreases CD133+ cancer stem cell population in colorectal cancer cell lines.

Citation Format: Sunghee Kim, Ja-Lok Ku. Metformin induces apoptotic cell death and inhibits CD133+ cancer stem cells in 5-Fu-resistant colorectal cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 780. doi:10.1158/1538-7445.AM2014-780