bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3947: Glutaminolysis inhibitors benzo[d][1,2]selenazol -3(2H)-one dimers completely disrupt mitochondrial function and showed early promise as effective antitumor agents with little general toxicity
Authors: Ruan Benfang Helen , Yu Yan , Chen Zhao , Jin Yanhong , Fang Jinzhang , Zhao Han , Ruan Jennifer
Published: August 2018

Targeting glutaminolysis is an emerging approach to solve the imminent problems of drug resistance and drug toxicity in cancer therapeutics. Ebselen was identified in the KGA coupled assay as a potent KGA inhibitor but showed very low potency in inhibiting the growth of glutamine - dependent cancer cells. For rigorous characterization, we developed a direct kinetic binding assay for KGA using Bio-Layer Interferometry (BLI) as the detection method; Ebselen was identified as a GDH inhibitor but not a KGA inhibitor. Proteomic analysis demonstrated that the ebselen derivative could crosslink with KGA protein, but the binding site is at the N-terminal appendix domain outside of the catalytic domain, which might explain why ebselen is not a potent KGA enzyme inhibitor in functional assays. Furthermore, we designed and synthesized several benzo[d][1,2]selenazol -3(2H)-one dimers which were subjected to SAR analysis by several glutaminolysis specific biochemical and cell based assays. Novel dual KGA/GDH inhibitors were discovered from the synthetic compounds; the dual inhibitors completely disrupt mitochondrial function, depleted cellular NAD(P)H levels, induced ROS mediated cell death, and inhibited the growth of various cancer cells but not the normal cells. Importantly, in an aggressive liver cancer mouse model, a KGA/GDH inhibitor reduced the tumor size by 50-60% and lowered plasma lactate dehydrogenase (a biomarker of tumor metastasis) by 2-3 times. Thus, the novel dual-specific KGA/GDH inhibitors showed early promise as effective antitumor agents with little general toxicity.

Citation Format: Benfang Helen Ruan, Yan Yu, Zhao Chen, Yanhong Jin, Jinzhang Fang, Han Zhao, Jennifer Ruan. Glutaminolysis inhibitors benzo[d][1,2]selenazol -3(2H)-one dimers completely disrupt mitochondrial function and showed early promise as effective antitumor agents with little general toxicity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3947.
Abstract 3950: Small molecule NSC59984 synergizes with ROS-generating agents to induce cell death via reduced GSH in cancer cells
Authors: Zhang Shengliang , Zhou Lanlan , El-Deiry Wafik S.
Published: August 2018

NSC59984 is a small molecule with a duel capability to induce mutant p53 degradation and restore p53 pathway signaling in cancer cells. NSC59984 requires reactive oxygen species (ROS) to induce mutant p53 degradation via ERK2. ROS regulates cell growth and death with tumor cells having higher ROS levels than normal cells. Increasing ROS level is a promising therapeutic strategy applied in cancer clinical trials. However, the toxicity of high ROS levels is a limitation that needs to be solved urgently. We investigated the synergy between NSC59984 and ROS in cancer cells. To increase ROS, cells were treated with BSO, an ROS-generating agent. Our data reveal that ROS sensitizes cells to NSC59984 treatment including enhanced-p53 restoration and mutant p53 degradation induced by NSC59984. Combination of NSC59984 and BSO shows a synergy to induce cell death in cancer cells with an antagonism observed in normal cells at the tested doses. Consistent with the in vitro experiments, our in vivo experiments show that the combination of NSC59984 and BSO significantly suppresses tumor growth. Immunohistochemical staining shows a decrease in Ki67 and an increase in cleaved caspase 3 in xenografted tumors following combination treatment, suggesting that the tumor suppression is due to the induction of cell death and the inhibition of cell growth. Furthermore, the combination of NSC59984 and BSO shows a significant reduction in glutathione (GSH) levels in cancer cells, suggesting that the synergy between NSC59984 and BSO is due to the inhibition of the antioxidant system in mutant p53-expressing cancer cells. Our data indicate that a high level of ROS can be considered as a biomarker for NSC59984 administration in cancer therapy and supports further evaluation of the combination of NSC59984 and ROS-generating agents.

Citation Format: Shengliang Zhang, Lanlan Zhou, Wafik S. El-Deiry. Small molecule NSC59984 synergizes with ROS-generating agents to induce cell death via reduced GSH in cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3950.
Abstract 3962: Antitumor effects of the antiparasitic agent ivermectin via inhibition of yes-associated protein 1 expression in gastric cancer
Authors: Kuramitsu Shotaro , Nanbara Sho , Hu Qingjiang , Kuroda Yousuke , Masuda Takaai , Ito Shuhei , Eguchi Hidetoshi , Mehara Yoshihiko , Mimori Koshi
Published: August 2018

[Background] Yes-associated protein 1 (YAP1) acts as an oncogene through dephosphorylation and nuclear translocation, and nuclear accumulation of YAP1 is associated with poor prognosis in gastric cancer (GC). We previously identified ivermectin, an antiparasitic drug, as a YAP1 inhibitor. Here, we aimed to clarify whether ivermectin had antitumor effects on GC through inhibition of YAP1.

[Result] First, we evaluated the antiproliferative effects of ivermectin on human GC cells using in vitro proliferation assays and a xenograft mouse model. YAP1-knockdown assays were performed to assess whether the sensitivity to ivermectin depended on YAP1 expression. Next, we explored the mechanism through which ivermectin regulated YAP1 expression or localization by immunoblotting and reverse transcription-quantitative polymerase chain reaction for YAP1 and the downstream gene CTGF. Finally, the clinical significance of YAP1 expression was examined using three independent GC datasets. We found that MKN1 GC cells were most sensitive to ivermectin, whereas MKN7 cells were most resistant. In MKN1 xenografts, ivermectin suppressed tumor growth, and the sensitivity of MKN1 cells to ivermectin was decreased by YAP1 knockdown. Ivermectin inhibited YAP1 nuclear expression and CTGF expression in MKN1 cells but not MKN7 cells. Moreover, ivermectin decreased YAP1 mRNA expression, thereby inhibiting nuclear accumulation of YAP1 in MKN1 cells. In survival analysis, low YAP1 mRNA expression was associated with a better prognosis in three independent GC datasets.

[Conclusion] Ivermectin suppressed the growth of GC in vitro and in vivo by inhibiting YAP1 expression. Furthermore, GC with low YAP1 expression had favorable clinicopathological features and good prognosis. These findings provided insights into the antiproliferative effects of ivermectin as a YAP1 inhibitor and established a theoretical basis for preclinical evaluations of ivermectin for management of GC.

Citation Format: Shotaro Kuramitsu, Sho Nanbara, Qingjiang Hu, Yousuke Kuroda, Takaai Masuda, Shuhei Ito, Hidetoshi Eguchi, Yoshihiko Mehara, Koshi Mimori. Antitumor effects of the antiparasitic agent ivermectin via inhibition of yes-associated protein 1 expression in gastric cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3962.
Abstract 3964: Mesothelin targeting immunotoxin LMB-100 alters the profile of tumor-secreted proteins
Authors: El-Behaedi Salma , Landsman Rebekah , Rudloff Michael , Kolyvas Emily , AlBalawy Rakan , Alewine Christine
Published: August 2018

Recombinant immunotoxins are antibody-based therapies that carry a bacterial toxin payload. LMB-100 is a next generation immunotoxin currently being tested in clinical trial. It targets the human cell surface antigen mesothelin (MSLN). MSLN is made by many solid tumors but has no expression in the parenchyma of vital organs. LMB-100 carries a Pseudomonas exotoxin payload that works by irreversibly modifying elongation factor-2 (EF-2), inactivating it and halting protein synthesis. To determine whether non-lethal doses of LMB-100 might inhibit tumor cell secretion of cytokines and growth factors that maintain the pancreatic cancer microenvironment, human pancreatic cancer cells were treated with LMB-100 and conditioned medium was assayed for tumor-secreted factors by ELISA and Luminex assays. Decreased levels of multiple secreted factors were detected including VEGF, PDGF, MMP-1, SPARC, and OPN. Conversely, levels of MIF, a stored cytokine released under stress conditions, increased dramatically upon LMB-100 treatment. To determine whether this also occurs in vivo, tumor-bearing athymic nude mice were treated with LMB-100 and levels of tumor-secreted factors in intratumoral fluid were measured. A statistically significant decrease in the same factors was found in treated mice compared to control mice. To determine whether these decreases in tumor-secreted factors could change the tumor microenvironment and alter the immune milieu, a mouse syngeneic model of pancreas cancer sensitive to LMB-100 was developed. LMB-100 treatment of this mouse cell model caused a statistically significant decrease in the angiogenesis factors VEGF and Angiopoietin-2 in conditioned medium. To determine if this also occurs in vivo, immune-competent mice bearing intraperitoneal or orthotopic tumors were treated with immunotoxin or vehicle then intratumoral fluid was assayed for secreted proteins contributed by both tumor and stromal cells. No detectable change in intratumoral fluid concentration was seen for VEGF, Antiopoietin-2 or a panel of 11 other secreted factors. In conclusion, LMB-100 does inhibit tumor cell production of secreted proteins, but this change is insufficient to alter the total concentration of secreted factors in the tumor microenvironment.

Citation Format: Salma El-Behaedi, Rebekah Landsman, Michael Rudloff, Emily Kolyvas, Rakan AlBalawy, Christine Alewine. Mesothelin targeting immunotoxin LMB-100 alters the profile of tumor-secreted proteins [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3964.
Abstract 3969: Lipocalin-2-targeted small interference RNA for inflammatory breast cancer treatment
Authors: Santiago-Sanchez Ginette S. , Valiyeva Fatma , Debeb Bisrat , Mejia Pablo E.
Published: August 2018

Inflammatory Breast Cancer (IBC) is an aggressive form of invasive breast cancer with low survival rate. Available therapies to treat this type of cancer are almost limited to surgery and radiation therapy. Potential therapeutic targets for IBC treatment include the oncogene Her2 and the Epidermal Growth Factor Receptor (EGFR). However, these therapies have not been successful since not all IBC tumors are Her2-amplified and development of resistance to treatment has also been observed. Hence, there is a critical need for novel therapeutic targets for IBC treatment. One of such possibilities is Lipocalin-2 (LNC2). LCN-2 is a secreted glycoprotein and an adipokine superfamily member that posses roles in transport of small lipophilic molecules in circulation, innate immunity, and iron homeostasis. High levels of LCN2 have been observed in invasive breast cancer human tissue. Additionally, LCN2 has been linked with breast cancer tumorigenesis and metastasis. Therefore, the purpose of our study is to assess if LCN2 is a plausible therapeutic target to treat IBC. Western blot analysis was used to assess the LCN2 protein level in a panel of IBC (MDA-IBC-3, SUM-149) and non-IBC cell lines (MCF-7, MDA-MB-231, MDA-MB-438). IBC cell lines showed a significantly higher expression of the LCN2 protein in comparison to the non-IBC cell lines. We decreased the expression of the LCN2 protein with small interference RNA (siRNA) molecules. Silencing of IBC cells with LCN2-targeted siRNA molecules decreased the LCN2 protein expression levels in almost fifty percent. Colony formation assay demonstrated that siRNA-mediated LCN2 silencing in MDA-IBC-3 and SUM-149 cell lines resulted in a significant decrease in cell proliferation. Taken together these results suggest that LCN2 may become a right target for IBC treatment. Ongoing experiments include cell invasion assays to determine if LCN2-targeted siRNAs reduce the metastatic potential, increase apoptosis and block cell cycle progression. Acknowledgements: This work is supported by the University of Puerto Rico Comprehensive Cancer Center and the U54 partnership with MD Anderson Cancer Research Center.

Citation Format: Ginette S. Santiago-Sanchez, Fatma Valiyeva, Bisrat Debeb, Pablo E. Mejia. Lipocalin-2-targeted small interference RNA for inflammatory breast cancer treatment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3969.
Abstract 3973: Dabrafenib suppresses the growth of BRAF-WT cancers through inhibition of novel targets Nek9 and Cdk16
Authors: Phadke Manali S. , Rix Lily , Smalley Inna , Bryant Annamarie , Lawrence Harshani , Schaible Braydon , Chen Ann , Rix Uwe , Smalley Keiran
Published: August 2018

Although the BRAF inhibitors dabrafenib and vemurafenib have both proven successful against BRAF-mutant melanoma, there seem to be differences in their mechanisms of action. Here, we show that dabrafenib is more effective at inhibiting the growth of NRAS-mutant and KRAS-mutant cancer cell lines than vemurafenib. Using mass spectrometry-based chemical proteomics we identified NEK9 and CDK16 as unique targets of dabrafenib. Both NEK9 and CDK16 were highly expressed in specimens of advanced melanoma, with high expression of both proteins correlating with a worse overall survival. A role for NEK9 in the growth of NRAS and KRAS-mutant cell lines was suggested by siRNA studies in which silencing was associated with decreased proliferation, cell cycle arrest associated with increased p21 expression, inhibition of phospho-CHK1, decreased CDK4 expression and the initiation of a senescence response. Inhibition of CDK4 but not CHK1 recapitulated the effects of NEK9 silencing, indicating this to be the likely mechanism of growth inhibition. We next turned our attention to CDK16 and found that its knockdown inhibited the phosphorylation of the Rb protein at S780 and increased expression of p27. Both of these effects were phenocopied in NRAS and KRAS-mutant cancer cells by dabrafenib but not vemurafenib. Combined silencing of NEK9 and CDK16 were associated with enhanced inhibition of melanoma cell proliferation and a greater induction of cell cycle arrest. In summary, we have identified dabrafenib as a potent inhibitor of NEK9 and CDK16, and suggest that inhibition of these kinases may have activity against cancers that do not harbor BRAF mutations.

Citation Format: Manali S. Phadke, Lily Rix, Inna Smalley, Annamarie Bryant, Harshani Lawrence, Braydon Schaible, Ann Chen, Uwe Rix, Keiran Smalley. Dabrafenib suppresses the growth of BRAF-WT cancers through inhibition of novel targets Nek9 and Cdk16 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3973.
Abstract 3975: Target engagement, thrombocytopenia, and efficacy induced by the dual Bcl2/xL inhibitor AZD4320 are quantitatively linked by a PK/PD model in leukemia xenografts
Authors: Gibbons Francis D. , Adam Ammar , Beaudoin Marie-Eve , Gangl Eric , Secrist Paul
Published: August 2018

The proteins Bcl2 and Bcl-xL are often up-regulated in cancer, and hold in check the apoptosis that would normally be initiated by accumulation of the BH3-only proteins Bax and Bak in response to genomic dysregulation. AZD4320 potently disrupts that interaction, initiating the apoptotic cascade in Bcl-2 or Bcl-xL-dependent tumors. Because platelets are known to be dependent on Bcl-xL, thrombocytopenia is an expected on-target effect. AZD4320 was administered at dose levels 0.5-10 mg/kg, both intravenously and extravascularly, to immune-compromised mice inoculated subcutaneously with the RS4;11 model of acute lymphocytic leukemia (ALL). Drug concentrations were measured by liquid chromatography-mass spectrometry (LC-MS) in plasma and tumor. A cleaved-caspase-3 ELISA was used to assess apoptotic activity in the tumor. Parallel efficacy studies, were used to assess tumor growth compared to vehicle, following tumors from initial regression at tolerated doses to regrowth. Tumors were measured using calipers, and tumor volumes computed using an ellipsoid approximation. We present a mini-physiologically based pharmacokinetic/pharmacodynamic (mPBPK/PD) model that links drug concentrations in plasma and tumor to observed caspase activity and efficacy in an integrated manner, across multiple dose levels and schedules. The tumor is modeled as a pool of sensitive cells which can be triggered rapidly by AZD4320 to apoptose, from which point they transition gradually to death, reducing tumor volume. Cleaved caspase-3 is used as a marker of apoptosis, and modeled using a sigmoidal response function with steep slope parameter. In this way, we effectively capture the transient nature of the response, despite AZD4320's long residence in the tumor. Thrombocytopenia is described not as an effect on the megakaryocyte precursors, but as a linear concentration-dependent effect on circulating platelets. Feedback from the circulation to megakaryocytes drives increased production to fill the deficit. Parameters are well-estimated throughout. Together, these components provide a comprehensive means to investigate the effects of dose and schedule with a dual Bcl2/BCL-xL inhibitor.

Citation Format: Francis D. Gibbons, Ammar Adam, Marie-Eve Beaudoin, Eric Gangl, Paul Secrist. Target engagement, thrombocytopenia, and efficacy induced by the dual Bcl2/xL inhibitor AZD4320 are quantitatively linked by a PK/PD model in leukemia xenografts [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3975.
Abstract 3979: Active components of Alpinia katsumadai-induced cell apoptosis and autophagy in human pancreatic cancer cells
Authors: An Weixiao , Lai Honglin , Zhang Yangyyang , Lin Xiukun , Cao Shousong
Published: August 2018

Alpinia katsumadai, as a traditional Chinese medicine, exhibited potent anti-oxidative and anticancer effects. Studies have shown that the effect of the active components of Alpinia katsumadai (ACAK) on anticancer is via apoptosis or autophagy in colorectal and glioblastoma cancer cells. However, the effects of ACAK on anticancer effect and inducation of apoptosis and autophagy in pancreatic cancer cells are still unknown. Therefore, we investigated the effects of ACAK on cytotocixity, apoptosis and autophagy as well as associated mechanisms in pancreatic cancers Panc-1 and Panc-28 cells. We found that ACAK showed greater inhibitory effects on the proliferation of Panc-1 and Panc-28 cells in a dose- and time-dependent manners (IC50 0.1692 and 0.1554mg/ml, respectively) compared to a variety of other cancer cells including breast cancer MDA-MB-468 cells, melamoma A875 cells, lung cancer A549 cells, colon cancer HCT-116 cells, and glioblastoma cancer U87 cells (IC50 0.2302, 0.2586, 0.2642, 0.3006, and 0.4298, respectively) by CCK-8 assay. Furthermore, ACAK induced cell apoptosis in pancreatic cancer Panc-1 and Panc-28 cells in a dose- dependent manner by Hochest 33342-PI and AnnexinV-PI dual staining analysis. Moreover, the expression levels of apoptosis related proteins casepase 3, casepase 9, and PARP were significantly up-regulated by Western blotting analysis. The study of cell autophagy was dectected by mRFP -GFP-LC3 adenovirus and western-blotting. and the results showed that the expression levels of autophagy-related protein beclin-1 and LC3-II were significanlt up-regulated by mRFP-GFP-LC3 adenovirus and western-blotting analysis. Our findings demonstrated that ACAK had greater cytotoxicity than a variety of other cancer cells by inducing apoptosis and autophagy in pancreatic cancer Panc-1 and Panc-28 cells. Therefore, ACAK may be developed as a novel anticancer agent for the treatment of pancreatic cancer clinically.

Citation Format: Weixiao An, Honglin Lai, Yangyyang Zhang, Xiukun Lin, Shousong Cao. Active components of Alpinia katsumadai-induced cell apoptosis and autophagy in human pancreatic cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3979.
Abstract 3981: Enhancement of the anticancer activity of methylenetetrahydrofolate dehydrogenase knockdown by folate depletion with carboxypeptidase G2
Authors: Rather Gulam M. , Kerrigan John E. , Scotto Kathleen W. , Bertino Joseph R.
Published: August 2018

The mitochondrial folate enzyme, methylene tetrahydrofolate dehydrogenase (MTHFD2), is a promising anticancer target because 1) it is required for the generation of one-carbon units required for purine synthesis and generation of NADH/NADPH necessary for protection from ROS in the mitochondria; 2) it is markedly distinct from the cytoplasmic MTHFD1 enzyme with respect to location, catalytic activity and cofactors; 3) it is expressed at low levels in proliferating normal cells; 4) in contrast, it is highly expressed in a variety of rapidly proliferating malignant tumors; 5) knockdown (KD) of MTHFD2 has been shown to inhibit proliferation in a subset of tumor cells in vitro; and 6) MTHFD2 KD is predicted to have metabolic consequences (glycine auxotrophy, folate deficiency) that could be exploited for combination therapy. In this study we tested the hypotheses that downregulation of MTHFD2 will result in relative folate deficiency, which can be exploited by using a folate-depleting enzyme, carboxypeptidase G2 (CPG2), to enhance tumor cell kill. CPG2 is approved by the FDA for the treatment of methotrexate overdose in patients. It acts to inactivate MTX or naturally occurring folates by hydrolyzing the terminal glutamate from folates and MTX, generating a pteroate and glutamate. As pteroates cannot be used to resynthesize folates, or MTX, this treatment depletes cells from MTX or folates. The combination of CPG2 in cell lines, MCF7 and T47D with MTHFD2 KD, was done and the actual increase in growth inhibition showed enhanced antitumor effects, indicating that when effective inhibitors of MTHFD2 are available, this combination may have widespread clinical usefulness, especially in rapidly proliferating tumors that express high levels of MTHFD2.

Citation Format: Gulam M. Rather, John E. Kerrigan, Kathleen W. Scotto, Joseph R. Bertino. Enhancement of the anticancer activity of methylenetetrahydrofolate dehydrogenase knockdown by folate depletion with carboxypeptidase G2 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3981.
Abstract 3982: Dual inhibition of HDAC and PI3 kinase signaling pathways with CUDC-907 in treatment of p53-driven medulloblastoma
Authors: Ronecker Jennifer S. , Pazniokas Julia , Amin Anubhav G. , Medeiros Raphael Salles Scortegagna de , Epelman Sidnei , Ghandi Chirag , Zanon Nelci , Schmidt Meic H. , Jhanwar-Uniyal Meena
Published: August 2018

Comprehensive studies of the medulloblastoma (MB) genome, epigenome and transcriptome have placed MB in four molecular subgroups: WNT, SHH, and two non-WNT/non-SHH: Group 3 and Group 4. Furthermore, studies suggest that p53-mutated SHH MBs frequently harbor genetic anomalies involving MYCN and GLI2 amplifications, which confer poor prognosis and resistance to treatments. The phosphatidylinositol-3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) intracellular signaling pathway that regulates cell growth, survival and invasion, is commonly deregulated in MB. Therefore, the PI3K/Akt/mTOR pathway may be a suitable target for therapeutic intervention. Moreover, Histone deacetylases (HDAC) regulate cancer initiation and progression, and studies have shown that small molecule inhibitors of HDAC can effectively limit MB proliferation. We aim to determine the status of tumor suppressor gene p53 in defining metastatic potential in SHH-MB and to establish the therapeutic efficacy of targeting these pathways in p53-driven MB cells with a dual agent or individual agents. Immunohistochemistry and FISH were used to determine the status of mutant p53 and MYC amplification or iso-p53, respectively, in MB tumors (n=41). p53-mutant MB cell line was used to investigate the signaling pathway that regulates proliferation, migration, and drug resistance using inhibitors of HDAC (LBH-589) and PI3K/mTOR (BKM-120/rapamycin) or CUDC-907 (dual inhibitor of HDACs and the PI3K/AKT). Results demonstrated that: 1) GAB-1 was highly expressed in the Shh group (82%) and KV1 expression was evenly distributed in all subtypes; 2) loss of p53 and overexpression of MYC varied in each subtype, but did not correlate with metastasis; 3) combined treatment with LBH-589 and BKM-120 or single treatment with CUDC-907 reduced cell proliferation, migration and S-phase entry; 4) MB cells were resistant to BKM -120, while LBH-589 or CUDC-907 caused apoptosis; 5) tumor formation was suppressed by BKM-120 given with mTOR inhibitors; 6) AKT and 4E-BP1 dephosphorylated following treatments with BKM-120 or CUDC-907. In conclusion, despite current clinical limitations, dual HDAC and PI3K inhibitors may benefit the understanding of aberrant signaling pathways in genetically driven MB, and may provide basis for future targeted therapies.

Citation Format: Jennifer S. Ronecker, Julia Pazniokas, Anubhav G. Amin, Raphael Salles Scortegagna de Medeiros, Sidnei Epelman, Chirag Ghandi, Nelci Zanon, Meic H. Schmidt, Meena Jhanwar-Uniyal. Dual inhibition of HDAC and PI3 kinase signaling pathways with CUDC-907 in treatment of p53-driven medulloblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3982.
Abstract 3986: HDAC4 degradation by combined TRAIL and valproic acid treatment induces apoptotic cell death of TRAIL-resistant head and neck cancer cells
Authors: Lee Bok-Soon , Kim Yeon Soo , Jang Jeon Yeob , Kim Hyo Jeong , Lee Myeong-Hoon , Chang Jae Won , Koo Bon Seok , Won Ho-Ryun , Kim Yong-Sung , Kim Chul-Ho
Published: August 2018

Purpose: Although TRAIL can directly induce death in cancer cells via DR4 and DR5 receptors, it appears that TRAIL resistance exists in many cancers. This study focuses on anti-cancer drugs for TRAIL-resistant head and neck cancer (HNC) to provide a further step toward effective cancer therapy.

Materials and Methods: The combined effects of TRAIL and VPA on cell viability were assessed via MTT and flow cytometric assays. Western blotting was used to determine cell death signaling.

Results: The results indicate that combined TRAIL and VPA treatment greatly reduced the cell viability and thus induced cell death relative to treatment with TRAIL or VPA alone in TRAIL-resistant HNC cells. A caspase-dependent signaling pathway was shown, and combined treatment with TRAIL and VPA also significantly decreased the expression of HDAC4. When we pretreated cells with z-VAD followed by combined treatment with TRAIL and VPA, the cell death was blocked with no decreased expression of HDAC4. To confirm the cell death involved in HDAC4 in HNC cells, we pretreated siRNA of the HDAC 4, followed by treating the TRAIL and VPA. The results showed that the knock down of HDAC4 sensitized the TRAIL-resistant HNC cells to apoptotic cell death. Finally, we showed a high expression of HDAC4 in HNC tissues compared to normal tissues obtained from the same patients.

Conclusion: Combined VPA and TRAIL treatment may be a promising therapy for HNC via HDAC4 degradation.

Citation Format: Bok-Soon Lee, Yeon Soo Kim, Jeon Yeob Jang, Hyo Jeong Kim, Myeong-Hoon Lee, Jae Won Chang, Bon Seok Koo, Ho-Ryun Won, Yong-Sung Kim, Chul-Ho Kim. HDAC4 degradation by combined TRAIL and valproic acid treatment induces apoptotic cell death of TRAIL-resistant head and neck cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3986.
Abstract 3993: Efficacy of SHP2 phosphatase inhibition in cancers with nucleotide-cycling oncogenic RAS, NF1 loss and RAS-GTP-dependent oncogenic BRAF
Authors: Bivona Trever G. , Nichols Robert , Haderk Franziska , Stahlhut Carlos , Schulze Christopher , Hemmati Golzar , Wildes David
Published: August 2018

Oncogenic alterations in the RAS-RAF-MEK-ERK pathway, including mutant forms of KRAS, BRAF, and loss of the tumor suppressor and RAS GTPase-activating protein (GAP) NF1, drive the growth of a wide spectrum of cancers. While BRAF and MEK inhibitors are efficacious against BRAFV600E-driven cancers, effective targeted therapies are lacking for most cancers driven by other pathway alterations, including oncogenic KRAS, NF1 loss, and non-V600E oncogenic BRAF. Here, we show that targeting the PTPN11/SHP2 phosphatase with a novel small molecule allosteric inhibitor is effective in human cancer models bearing nucleotide-cycling oncogenic RAS (e.g. KRASG12C), NF1 loss, or RAS-GTP dependent oncogenic BRAF (e.g. class 3 BRAF mutants). SHP2 inhibitor treatment decreases oncogenic RAS-RAF-MEK-ERK signaling and cancer growth by disrupting SOS1-mediated RAS-GTP loading. Our findings illuminate a critical function for SHP2 in promoting oncogenic RAS activation and downstream signaling in cancers with nucleotide-cycling oncogenic RAS, RAS-GTP dependent oncogenic BRAF, and NF1 loss. SHP2 inhibition is a promising molecular therapeutic strategy for patients with cancers bearing these oncogenic drivers.

Citation Format: Trever G. Bivona, Robert Nichols, Franziska Haderk, Carlos Stahlhut,Christopher Schulze, Golzar Hemmati, David Wildes. Efficacy of SHP2 phosphatase inhibition in cancers with nucleotide-cycling oncogenic RAS, NF1 loss and RAS-GTP-dependent oncogenic BRAF [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3993.
Abstract 3998: Integrative molecular analysis uncovers key molecules and signaling pathways regulated by RKIP in gastrointestinal stromal tumors (GISTs)
Authors: Campanella Nathália C. , Melendez Matias , Leal Leticia Ferro , Evangelista Adriane Feijo , Faça Vitor Marcel , Reis Rui Manuel
Published: August 2018

Introduction: Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal tumor on the gastrointestinal tract, and are characterized by hotspot mutations in KIT and PDGFRA genes, which are predictive of imatinib-based therapy response. We showed previously that RKIP, a metastasis suppressor gene, is lost in about 17% of GIST, and it was associated with an adverse outcome. RKIP is plays a pivotal role in several signaling pathways, namely MAPK, NF-κβ, and GSK3β however a multidimensional integrative analysis to uncover its biological impact was never done in cancer and in particular in GIST.

Experimental procedures: We established a RKIP knockout GIST cell line (GIST-T1 KO) by CRISPR/Cas9 system. The differential gene and microRNA expression profile between GIST-T1 RKIP-KO and GIST-T1 negative control cells were assessed using the NanoString nCounter® technology and the nCounter® PanCancer Pathways and nCounter® miRNA Expression assays (NanoString Technologies). All procedures including sample preparation (100 ng of RNA), hybridization, detection and scanning were performed according to manufacture's instructions (NanoString Technologies). In a second step, we performed a global proteomic analysis using the Acquity UPLC M-Class chromatography system (Waters Co) linked to the mass spectrometer Synapt G2-Si (Waters Co). The protein

identification and quantification were performed in Progenesis QI for Proteomics (Waters Co) software.

Results: The nCounter gene expression analysis showed 56 genes differentially expressed: 18 downregulated and 38 upregulated genes in RKIP KO cells. The altered genes are especially involved in PI3k-AKT, MAPK and Ras signaling pathways. The global proteomic characterization showed 50 proteins differentially expressed: 5 were downregulated and 44 upregulated proteins in RKIP KO cells. In microRNA expression profile analysis we found 133 downregulated microRNAs in RKIP KO cells. Taking together, these findings uncover new molecules that are going to screen in a cohort of GIST patients.

Conclusion: For the first time, using a multidimensional integrative analysis we identified driver candidates and pathways, which will contribute the uncovered the biological role of RKIP in cancer.

Citation Format: Nathália C. Campanella, Matias Melendez, Leticia Ferro Leal, Adriane Feijo Evangelista, Vitor Marcel Faça, Rui Manuel Reis. Integrative molecular analysis uncovers key molecules and signaling pathways regulated by RKIP in gastrointestinal stromal tumors (GISTs) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3998.
Abstract 40: The role of pressure-driven flow in invasion and chemoresistance of cancer cells in an engineered breast tumor model
Authors: Simi Allison K. , Kourouklis Andreas P. , Piotrowski-Daspit Alexandra S. , Tien Joe , Nelson Celeste M.
Published: August 2018

Collapsed blood or lymphatic vessels in the tumor microenvironment often cause fluid buildup, leading to heterogeneous flow throughout the tissue. Here, we used a three-dimensional (3D) engineered tumor model to investigate how fluid flow specifically influences invasion and chemoresistance of breast cancer cells. To mimic breast tumors, we cultured aggregates of MDA-MB-231 human breast cancer cells embedded in 3D collagen channels. Collagen channels were flanked on the ends by two media reservoirs. By changing the relative heights of media in the reservoirs, we controlled the pressure-induced flow experienced by the tumor cell aggregate. We found that the direction of flow through the collagen channel determined the invasive phenotype of the engineered tumor. These analyses will be repeated with the addition of chemotherapy drugs taxol or 5-fluorouracil in the media to determine the effect of fluid flow on chemotherapeutic response. Our engineered tumor model provides insight into how physical forces influence the invasive phenotype of cancer cells.

Citation Format: Allison K. Simi, Andreas P. Kourouklis, Alexandra S. Piotrowski-Daspit, Joe Tien, Celeste M. Nelson. The role of pressure-driven flow in invasion and chemoresistance of cancer cells in an engineered breast tumor model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 40.
Abstract 400: Noncanonical role of SEC23B in thyroid carcinogenesis: Anemia gene meets cancer
Authors: Yehia Lamis , Ni Ying , Jindal Supriya , Abbas Ata , Komar Anton A. , Eng Charis
Published: August 2018

Hereditary cancer syndromes serve as powerful models to uncover cancer-relevant genes. Cowden syndrome (CS), typically caused by germline PTEN mutations, is a heritable cancer syndrome with high risks of breast and thyroid cancers. However, there is a growing subset of CS without PTEN mutations. By exome sequencing cum family-studies, we identified heterozygous SEC23B variants in CS-associated and apparently sporadic thyroid cancer. Functional characterization revealed mutant CS-associated SEC23B led to endoplasmic reticulum (ER)-stress-mediated cell-colony formation and survival, growth and invasion, reflecting major hallmarks of cancer.

Classically, SEC23B is a component of coat protein complex II (COPII) vesicles that transport secretory proteins from the ER to the Golgi apparatus. Germline homozygous or compound heterozygous loss-of-function SEC23B mutations cause Congenital Dyserythropoietic Anemia Type II (CDA II), resulting in decreased SEC23B levels. Intriguingly, our in vitro data does not recapitulate the CDA II-like decreased SEC23B protein expression, suggesting possible change-of-function effects in a cancer context. Here, we show mutant SEC23B exists within nucleoli, besides classical distribution at the ER/Golgi. This occurs in the absence of nucleolar localization of other COPII protein components and does not compromise canonical secretory function. Global transcriptomic and polysome profiling analyses reveal increased expression of ribosomal protein and translation-related genes, accompanied by enhanced translational capacity in mutant cells in response to ER stress. We also show mutant SEC23B directly binds to Upstream Binding Transcription Factor (UBF), concomitantly with increased UBF binding at the ribosomal DNA (rDNA) promoter region with ER stress. Unbiased proteomic analyses further show SEC23B, both wildtype and mutant, can interact with proteins in the ER stress and EIF2 signaling pathways. We validate through immunoprecipitation the binding interaction between SEC23B and UBA52, with notably increased binding with mutant SEC23B. UBA52 is ribosomal protein RPL40, known to regulate selective translation of stress-related transcripts, protein synthesis, and the cell cycle. Knockdown of SEC23B in vitro resulted in a concomitant decrease in protein levels of UBA52, further supporting that SEC23B and UBA52 are potentially co-regulated. All in all, our data suggest that SEC23B has non-canonical COPII-independent function, particularly within the ribosome biogenesis pathway, that may contribute to the pathogenesis of cancer-predisposition.

Citation Format: Lamis Yehia, Ying Ni, Supriya Jindal, Ata Abbas, Anton A. Komar, Charis Eng. Noncanonical role of SEC23B in thyroid carcinogenesis: Anemia gene meets cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 400.
Abstract 4002: EGR1 downregulation by miR-377-3p mediated lung tumorgenesis following benzo[a]pyrene exposure
Authors: Ke Xinxin , Shen Jing , Shao Jimin , Qi Hongyan
Published: August 2018

Polycyclic aromatic hydrocarbons (PAHs), especially Benzo[a]pyrene (B[a]P), contained in cigarette smoke and air pollution, belong to a category of important carcinogens for lung cancer. B[a]p exerts its carcinogenic function by causing mutations, cytotoxicity and inhibiting DNA synthesis. However, the early molecular events and related mechanisms of B[a]P-induced cell transformation and tumor initiation remains unclear. In this study, we showed that EGR1 was significantly downregulated during human bronchial epithelial BEAS-2B cell transformation and mice lung carcinogenesis after B[a]P and its active form B[a]PDE exposures, respectively. Using RNA-seq to screening and using qRT-PCR, western-blot and immunohistochemistry for validation. In contrast, overexpression of EGR1 inhibited the cell malignant transformation upon B[a]PDE exposure. Furthermore, miR377-3p was strongly increased by B[a]PDE/B[a]P in vitro and in vivo. And luciferase reporter assays showed that the exposure-induced miR377-3p was crucial for inhibition of EGR1 by targeting its 3'-UTR. Importantly, miR-377-3p antagomir reversed the effect of EGR1 downregulation in the cell malignant transformation and tumor initiation models. The target genes of EGR1, ANKRD1 and ATF3, which could be involved in B[a]P-induced lung tumorgenesis, were also analyzed in cell and mice model, respectively. Finally, the B[a]P exposure-induced molecular changes were examined by immunohistochemistry in clinical lung cancer tissues and evaluated with TCGA data. Taken together, the results demonstrated that Benzo[a]pyrene exposure may induce lung tumorgenesis through miR-377-3p mediated reduction of EGR1 expression, suggesting an important role of the tumor suppressor EGR1 in PAH-induced lung carcinogenesis.

Keywords: EGR1, miR-377-3p, malignant transformation, lung tumorgenesis, gene regulation

Citation Format: Xinxin Ke, Jing Shen, Jimin Shao, Hongyan Qi. EGR1 downregulation by miR-377-3p mediated lung tumorgenesis following benzo[a]pyrene exposure [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4002.
Abstract 4006: Serine protease PRSS8 suppresses colorectal carcinogenesis and metastasis
Authors: Bao Yonghua , Guo Yongchen , Yang Yiqiong , Wei Xiaonan , Zhu Xiangdong , Zhang Wei , Yang Wancai
Published: August 2018

PRSS8 is a serine protease and has important physiological and pathophysiological functions, but its roles in cancer initiation and progression are unclear. In this study, a conditional knockout mouse model of Prss8 fl/fl,p-Villin-Cre+ mice were developed and thoroughly characterized at the ages of 3, 6 and 9 months. Intestinal epithelial cell proliferation, migration, and differentiation, were determined. The approaches of gain- and loss of expression of PRSS8 in human colon cancer cells were performed to evaluate anti-cancer and anti-metastasis in vitro and in nude mice by mouse subcutaneous injection, tail vein injection and intra-spleen transplantation. Gene profile and gene set enrichment analysis and mechanistic studies were conducted in vitro and in human colorectal cancer tissues. Results: Genetic deficiency of the Prss8 gene caused spontaneous formation of colitis and inflamed rectum at the early age, but caused intestinal tumors and lymph node metastasis at late age, which was linked to increased intestinal cell proliferation and migration, but decreased cell differentiation. Increased PRSS8 expression showed inhibition of cancer cell growth and metastasis in nude mice, and inhibited migration, invasion, colony formation and tumor sphere formation in vitro, but reduced expression of PRSS8 facilitated malignancies. Mechanistic studies revealed that PRSS8 targets Wnt/β-catenin, epithelial-mesenchymal-transition, and stem cell signaling pathways, which was supported by the results from the Cancer Genome Atlas, and were validated in human colorectal cancers. Conclusion: PRSS8 is a novel tumor suppressor that plays critical roles in the suppression of carcinogenesis and metastasis via targeting Wnt/β-catenin, EMT and stem cell signaling pathway.

Citation Format: Yonghua Bao, Yongchen Guo, Yiqiong Yang, Xiaonan Wei, Xiangdong Zhu, Wei Zhang, Wancai Yang. Serine protease PRSS8 suppresses colorectal carcinogenesis and metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4006.
Abstract 4014: APOBEC3B promotes hepatocarcinogenesis and metastasis through novel deaminase-independent activity
Authors: Ng Irene O L , Ma Wei , Ho Daniel WH , Sze Karen MF , Tsui Yue M. , Chan Lo K. , Lee Joyce MF
Published: August 2018

APOBEC3B (A3B), a cytidine deaminase, is known to play important roles in creating de novo genomic C-to-T mutations in cancers and contribute to induction of genomic instability. In this study, we evaluated the roles of A3B in the progression and metastasis of human hepatocellular carcinoma (HCC). Using whole-transcriptome and whole-exome sequencing and quantitative real-time PCR, we found that A3B was overexpressed in human HCCs and A3B expression was significantly correlated with the proportion of genomic C-to-A and G-to-T mutations. Upon clinicopathological correlation, higher A3B expression was associated with more aggressive tumor behavior. Wild-type A3B (wt-A3B) overexpression in HCC cells promoted cell proliferation, and cell migratory and invasive abilities in vitro, and tumorigenicity and metastasis in vivo. On the other hand, knockdown of A3B suppressed cell proliferation, migratory and invasive abilities of HCC cells with high endogenous A3B level. However, to our surprise, overexpression of A3B deaminase-dead double mutant (E68A/E255Q) led to similar results as wt-A3B in HCC both in vitro and in vivo. Furthermore, overexpression of wt-A3B and mutant A3B both enhanced cell cycle progression in HCC cells. Altogether, our data demonstrated a novel deaminase-independent role of A3B in contributing to HCC tumorigenesis and metastasis.

Citation Format: Irene O L Ng, Wei Ma, Daniel WH Ho, Karen MF Sze, Yue M. Tsui, Lo K. Chan, Joyce MF Lee. APOBEC3B promotes hepatocarcinogenesis and metastasis through novel deaminase-independent activity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4014.