bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 703: An anthelmintic drug, flubendazole, exerts antitumor effects in triple-negative breast cancer via targeting cancer stem-like properties
Authors: Kim Yoon-Jae , Oh Eunhye , Cho Tae-Min , Jang Seojin , Park Jung Min , Park Soeun , Park Minsu , Kim Ji Young , Seo Jae Hong
Published: August 2019

Triple-negative breast cancer (TNBC) remains challenging due to the absence of established molecular targets for the phenotype. These tumors tend to be more aggressive with a higher risk of recurrence and metastasis, leading to poorer clinical outcomes. Flubendazole (FLU) is a widely used anthelmintic agent that also exhibits anticancer activity in several cancer types. The objective of the present study was to investigate the effects of FLU on cell proliferation, apoptosis and breast cancer stem cell (BCSC)-like properties in TNBC. The effect of FLU on TNBC cell lines in vitro was evaluated in terms of apoptosis, cell cycle distribution, caspase-3/-7 activity and BCSC-like characteristics. An orthotopic allograft model with 4T1 mammospheres was used to examine the effect of FLU on tumor growth and angiogenesis in vivo. FLU efficiently induced apoptosis with accompanied by activation of caspase-3 and -7, and PARP cleavage in TNBC. FLU-induced apoptosis is associated with the accumulation of G2/M phase cells by tubulin degradation, leading to mitotic catastrophe. Interestingly, FLU significantly suppressed BCSC-like properties in TNBC, as evidenced by a marked inhibition of ALDH1 activity, the CD44high/CD24low and CD49fhigh/CD24high subpopulations, and the subsequent impairment of mammosphere formation in vitro. This appears to contribute toward the suppression of BCSC-enriched tumor burden and intra-tumoral angiogenesis in TNBC allografts, together with a significant downregulation of ALDH1A1 and CD49f expression in vivo. Our findings suggest that FLU kills not only rapid proliferating tumor cells but also effectively eradicates BCSC-like cells in vitro and in vivo. These findings warrant further investigation of flubendazole as a potential treatment for triple-negative breast cancer.

Citation Format: Yoon-Jae Kim, Eunhye Oh, Tae-Min Cho, Seojin Jang, Jung Min Park, Soeun Park, Minsu Park, Ji Young Kim, Jae Hong Seo. An anthelmintic drug, flubendazole, exerts antitumor effects in triple-negative breast cancer via targeting cancer stem-like properties [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 703.
Abstract 704: RALB GTPase: A critical regulator of DR5 cell surface expression and TRAIL sensitivity in RAS mutant colorectal cancer
Authors: Khawaja Hajrah , Roberts Jamie Z. , Javadi Arman , O'Reilly Paul , McArt Darragh , Allen Wendy L. , Morrison Markus , Kennedy Richard , Vitale Nicolas , Johnston Patrick G. , Harrison Tim , Longley Daniel B. , Evergren Emma , Schaeybroeck Sandra Van
Published: August 2019

RAS mutant (MT) metastatic colorectal cancer (mCRC) remains a difficult-to-treat group. RALA and RALB GTPases function downstream of RAS and have been found to be key regulators of several cell functions and implicated in KRAS-driven tumourigenesis. However, their role as regulators of the apoptotic machinery remains to be elucidated. Here, we found that inhibition of RALB expression, but not RALA, resulted in caspase 8-dependent cell death in KRASMT cells, that was not further increased following MEK1/2 inhibition. Proteomics analysis and mechanistic studies revealed that RALB inhibition induces marked upregulation of the pro-apoptotic cell-surface TRAIL Death Receptor 5 (DR5) receptor in KRASMT CRC. Moreover, DR5 knockout also attenuated siRALB-induced apoptosis, confirming the role of the extrinsic apoptotic pathway as regulator of siRALB-induced cell death. Importantly, TRAIL treatment resulted in acute association of RALB with the death-inducing signalling complex (DISC) and targeting RALB using pharmacologic inhibition or RNAi approaches, resulted in a potent increase in TRAIL-induced cell death in RASMT CRC. Significantly, high RALB mRNA levels were found in the poor prognostic CRIS-B CRC subgroup. Collectively, this study provides the first evidence for a role of RALB in apoptotic priming and that RALB inhibition may be a promising strategy to improve response to TRAIL treatment in poor prognostic RASMT CRC.

Citation Format: Hajrah Khawaja, Jamie Z. Roberts, Arman Javadi, Paul O'Reilly, Darragh McArt, Wendy L. Allen, Markus Morrison, Richard Kennedy, Nicolas Vitale, Patrick G. Johnston, Tim Harrison, Daniel B. Longley, Emma Evergren, Sandra Van Schaeybroeck. RALB GTPase: A critical regulator of DR5 cell surface expression and TRAIL sensitivity in RAS mutant colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 704.
Abstract 707: Emerging role of mitochondrial pathway in crosstalk between ferroptosis and apoptosis
Authors: Lee Yong J.
Published: August 2019

Introduction: Several processes lead to cell death; ferroptosis, apoptosis, necrosis, and autophagy are the primary mechanisms. Each type of biological death has been believed to have a distinct biochemical and morphological fingerprint. However, emerging evidence suggests that these different types of cell death often share common pathways. In this study, we investigated whether mitochondrial signaling pathways play an important role in the intracellular communication between ferroptosis and apoptosis.

Methods: The human colorectal carcinoma HCT116 cell line was used to evaluate the interplay between cell death pathways. Cells were treated with the apoptotic agent TRAIL (tumor necrosis factor-related apoptosis-inducing ligand) and ferroptotic agents such as artesunate and erastin. Cell death was determined by trypan blue exclusion assay. Apoptosis was measured by western blot. Ferroptosis was assessed using MDA assay. Bax oligomerization was performed by mitochondrial fractionation followed by SDS-PAGE under nondenaturing conditions.

Results: We observed that the combinatorial treatment induced synergistic apoptosis, which was mediated through an increase in caspase activation. We also observed an increase in pro-apoptotic Bax oligomerization in the mitochondrial fraction. The combined treatment-induced apoptosis was suppressed in Bax-deficient HCT116 cells. Ferroptotic agents enhanced the intracellular level of anti-apoptotic Mcl-1. However, the combined treatment decreased the level of Mcl-1. Interestingly, the combined treatment-induced apoptosis was abolished in HCT116 cells with knockin of Mcl-1 phosphorylation site mutant (S121A/E125A/S159A/T163A).

Conclusions: These data suggest a novel mechanism for the combined treatment of the apoptotic agent TRAIL and ferroptotic agent erastin/artesunate-induced synergistic apoptosis through the mitochondrial pathway.

Citation Format: Yong J. Lee. Emerging role of mitochondrial pathway in crosstalk between ferroptosis and apoptosis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 707.
Abstract 708: Targeting fatty acid synthase induces apoptosis and senescence
Authors: Steen Travis Van der , Kemble George , Lupu Ruth
Published: August 2019

Fatty acid synthase (FASN) is a novel therapeutic target for cancer as it has increased expression in many different types of cancers. There has been significant understanding that cancer cells can hijack and modify tightly regulated metabolic pathway for its own survival. FASN is the enzyme that synthesizes palmitate from malonyl-CoA for energy storage, structure and biomolecules. FASN expression is low to undetectable in normal cells with the exception of liver and adipose tissue. In this study, we investigated the role of FASN in breast cancer cells. We observed that in highly expressing FASN breast cancer cells, TVB-3166, FASN inhibitor, results in a decrease in cellular proliferation and an increase in cellular death. FASN inhibition increased reactive oxygen species (ROS) and altered NADPH/NADP+ ratio. Increase in ROS and NADPH have been linked to the intrinsic apoptosis pathway. Additionally, there is an increase in expression of the BH3-only proteins Bim, Puma and Noxa, leading to the permeabilization of the mitochondria and the release of cytochrome c. Furthermore, after prolong exposure to TVB-3166 a number of cells enter into a senescent state. This was observed the in detection of β-galactosidase and confirmed by the expression of senescent markers including ARF, INK4A, and CIP. Additionally, when the cells were treated with a CDK4/6 inhibitor in combination with the FASN inhibitor TVB-3166, an increase in stress induced premature senescence was observed. Our data demonstrates that inhibition of FASN induces intrinsic apoptosis while it stimulates cellular senescence to the apoptotic resistant cells. Therefore, the ability for TVB-3166 to both eradicate and overturn cell growth makes it potential therapeutic modality.

Citation Format: Travis Van der Steen, George Kemble, Ruth Lupu. Targeting fatty acid synthase induces apoptosis and senescence [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 708.
Abstract 713: BBI-608 modulates DNA repair pathways, induces apoptosis and impacts response to 5-Fluorouracil and ionizing radiation in pancreatic cancer cells
Authors: Farran Batoul , Nagaraju Ganji P. , Shipra Sneha G. , Bethi Shipra R. , Lesinski Gregory B. , El-Rayes Bassel
Published: August 2019

Background: BBI-608 has been shown to modulate multiple oncogenic cellular pathways which are often constitutively active in pancreatic ductal adenocarcinoma (PDAC). Among these are key mediators of DNA repair and cellular survival pathways. We hypothesized that BBI-608 would sensitize PDAC cells to chemoradiotherapy.

Methods: The combined effects of BBI-608 and chemoradiotherapy (5-FU + IR) were evaluated in human (MIA PaCa-2) and murine (PANC-02) PDAC cell lines using a Br-dU cell proliferation assay. To elucidate the mechanism of action, effects on the expression of DNA damage and repair (pATM, γ-H2AX, p53; pATR, and Rad51) molecules were examined by Western blot. Apoptosis was assessed via Annexin V staining and flow cytometric analysis to evaluate mechanism of cell death following treatment of MIA PaCa-2 and PANC-02 cell lines. The in vivo activity of BBI-608, 5-FU + IR, alone and in combination, were evaluated in C57BL/6 mice bearing subcutaneous PANC-02 tumors.

Results: The combination of BBI-608 and chemoradiotherapy significantly decreased (p<0.001) cell proliferation and induced Annexin V positive cells in both MIA PaCa-2 and PANC-02 cell lines. BBI-608 and chemoradiotherapy also decreased the activation of pATR, MDM2, and Rad51 and increased pATM, γ-H2AX and p53. Administration of BBI-608 potentiated the effects of chemoradiotherapy in vivo in mice bearing subcutaneous PANC-02 tumors as measured by tumor volume.

Conclusion: These data identify a role for BBI-608 as a modulator of DNA damage and apoptosis. These effects enhanced the efficacy of chemoradiotherapy in cell lines and in vivo models of PDAC and provide rationale for continued investigation in the pre-clinical and clinical setting.

Note: This abstract was not presented at the meeting.

Citation Format: Batoul Farran, Ganji P. Nagaraju, Sneha G. Shipra, Shipra R. Bethi, Gregory B. Lesinski, Bassel El-Rayes. BBI-608 modulates DNA repair pathways, induces apoptosis and impacts response to 5-Fluorouracil and ionizing radiation in pancreatic cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 713.
Abstract 744: Clinically targetable genomic alterations in acral melanoma
Authors: Broit Natasa , Dutton-Regester Ken , Johansson Peter , Pritchard Antonia L. , Boyle Glen M. , Hayward Nicholas K.
Published: August 2019

Background: Acral melanoma is a rare subtype of melanoma, which has a distinct genomic profile from cutaneous melanoma. While survival outcomes for late-stage cutaneous melanoma have significantly improved in the last decade, treatment options for acral melanoma remain limited. The purpose of this study was to assess therapeutically targetable genomic alterations in acral melanoma.

Methods: The AACR project GENIE database was accessed via cBioPortal (http://cbioportal.org/genie/; version 3) to identify recurrent gene alterations in acral melanomas analyzed using the MSK-IMPACT oncopanel. These alterations were assessed through Cancer Genome Interpreter (http://cancergenomeinterpreter.org) to identify clinically actionable alterations.

Results: Thirty-six patients with acral melanoma were identified from the cBioPortal-GENIE database. These patients were sequenced using three different versions of oncopanel, which included 341 (n = 6), 410 (n = 22) and 468 (n = 8) genes. Seventy-six genes were found to be somatically mutated in this cohort in at least one sample. The most commonly mutated genes were NF1 (n = 5), BRAF (n = 5), PTPRT (n = 4), NOTCH3, HRAS and KRAS (n = 3). Copy number variations greatly contributed to aberration burden, with 132 unique genes found to either carry an amplification or deletion event. The most commonly amplified genes were CDK4 (n = 9), CCND1 (n = 9), FGF19/FGF4 (n = 8), PAK1 (n = 8), MDM2 (n = 7) and FGF3 (n = 7). The most recurrently deleted genes included CDKN2A (n = 9) and CDKN2B (n = 8), and less commonly JAK2 and PTEN (n = 2). Some genes were altered by various mechanisms; for example, some tumors had KIT activated by amplification while other tumors carried missense mutations. BRAF exhibited two fusion events: BRAF-METTL2B (with concomitant amplification) and BRAF-KIAA1549. Cancer Genome Interpreter identified several therapeutic targets, some of which have been the focus of clinical trials in cutaneous melanoma and other solid tumors, e.g. CDK4/6 inhibitors to target amplified CDK4 and CCND1, or mTOR inhibitors for tumors with NF1 mutations.

Conclusion: Several inhibitors are available which could show efficacy in a subset of acral melanomas with particular genomic alterations. Many of these inhibitors have yet to be tested in the context of acral melanoma and should be explored.

Citation Format: Natasa Broit, Ken Dutton-Regester, Peter Johansson, Antonia L. Pritchard, Glen M. Boyle, Nicholas K. Hayward. Clinically targetable genomic alterations in acral melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 744.
Abstract 75: Anti-tumor activity of a novel selenonucleoside via targeting Skp2 degradation in paclitaxel-resistant prostate cancer
Authors: Byun Woong Sub , Jin Minkyung , Yu Jinha , Kim Won Kyung , Jeong Lak Shin , Lee Sang Kook
Published: August 2019

Prostate cancer (PC) is the most common disease in men over age 50, and its prevalence rate has been gradually increasing since 1980. Taxane-derived anticancer agents are the primary agents used to treat metastatic prostate cancer patients; however, the side effects and acquired drug resistance limit the success of these therapies. Because there is no specific treatment for paclitaxel-resistant prostate cancer, it is necessary to develop new targets and therapeutic strategies to overcome the acquired resistance. In this study, the antitumor activity of a novel selenonucleoside (4′-selenofuranosyl-2,6-dichloropurine, LJ-2618), a third-generation nucleoside, and its plausible mechanisms of action in paclitaxel-resistant prostate cancer (PC-3-Pa) cells were investigated. The established PC-3-Pa cells exhibited over 100-fold resistance against paclitaxel compared to the paclitaxel-sensitive PC-3 cells. LJ-2618, however, effectively inhibited the proliferation of both cell lines with similar IC50values in vitro. In PC-3-Pa cells, the activated PI3K/Akt signaling pathway was suppressed by LJ-2618 treatment. In addition, Skp2 was found to be over-expressed in paclitaxel-resistant cells, and the transfection of Skp2 siRNA recovered the sensitivity of paclitaxel in PC-3-Pa cells. Furthermore, LJ-2618 significantly down-regulated Skp2 expression in PC-3-Pa cells by promoting degradation and inducing destabilization of Skp2, which triggers G2/M cell cycle arrest. In a xenograft mouse model implanted with PC-3-Pa cells, LJ-2618 (3 or 10 mg/kg) effectively inhibited tumor growth with the enhancement of Skp2 degradation and induction of p27 expression in tumor tissues. These findings suggest that LJ-2618 may have a potential for overcoming paclitaxel resistance via promoting Skp2 degradation and stabilizing p27 expression in PC-3-Pa cells. Therefore, the novel selenonucleoside LJ-2618 may lead to the development of a new treatment strategy for patients with paclitaxel-resistant, castration-resistant prostate cancer. This study was supported by a National Research Foundation of Korea (NRF) Grant funded by the Korean Government (NRF-2016M3A9B6903499).

Citation Format: Woong Sub Byun, Minkyung Jin, Jinha Yu, Won Kyung Kim, Lak Shin Jeong, Sang Kook Lee. Anti-tumor activity of a novel selenonucleoside via targeting Skp2 degradation in paclitaxel-resistant prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 75.
Abstract 753: In situ RNA expression profiling of 1600+ immuno-oncology targets in FFPE tissue using NanoString GeoMx™Digital Spatial Profiler
Authors: Hoang Margaret , Zhou Zoey , Kriner Michelle , Sorg Kristina , Norgaard Zach , Piazza Erin , Merritt Chris , Kim Dae , Beechem Joseph
Published: August 2019

Clinical specimens including formalin-fixed, paraffin embedded (FFPE) tumor sections preserve spatial and molecular information of tumor cells and their surrounding microenvironment. This valuable spatial information is loss with bulk RNA-seq, the most prevalent method for gene expression profiling of archival FFPE samples. In contrast, the NanoString GeoMx™ Digital Spatial Profiler (DSP) is a high multiplexing assay that can profile thousands of RNAs from user selectable regions of interest (ROIs) in FFPE sections. Here we introduce a 1600+ gene panel of tumor, stroma, and immune cell-specific content derived from NanoString nCounter® PanCancer series. We compare RNA profiling using our 1600+ immune-oncology panel to the RNA-seq methodology in FFPE samples. DSP RNA in situ probes are photocleavable oligonucleotides tags coupled to hybridization sequences that bind to mRNA transcripts in the FFPE tissue section. We gridded 96 ROIs upon the FFPE section, photocleaved oligonucleotide tags from each ROI were collected and prepared into sequencing libraries with our NGS readout workflow. After sequencing, reads were charted back to each ROI in the tissue section, generating a map of transcript activity within the tissue. We found high concordance between “bulk” DSP RNA (counts from all 96 gridded ROIs) and RNA-seq from the same FFPE block. Individually, however, each ROI showed different expression patterns than bulk and ROI expression patterns clustered based on similar tissue morphology. We further profiled the tumor and microenvironment compartments from multiple FFPE cancer tissues, comparing our 1600+ RNA expression profile to RNA-seq. DSP was able to detect a higher number of genes with low expression within each spatial compartment compared to bulk RNA-seq. These data demonstrate that DSP offers unparalleled sensitivity for large-scale gene expression while preserving critical information about tissue architecture. GeoMx™ DSP technology is for Research Use Only and not for use in diagnostic procedures.

Citation Format: Margaret Hoang, Zoey Zhou, Michelle Kriner, Kristina Sorg, Zach Norgaard, Erin Piazza, Chris Merritt, Dae Kim, Joseph Beechem. In situ RNA expression profiling of 1600+ immuno-oncology targets in FFPE tissue using NanoString GeoMx™Digital Spatial Profiler [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 753.
Abstract 754: A comprehensive immunogenomics profiling platform enables the exploration of the intricate relationship between TMB, MSI, Neoantigen and HLA status among multiple cancers
Authors: Wang Jie , Li Ao , Wang Jiawei , Cheng Zhaoze
Published: August 2019

With the rapid advancement of immune therapy on cancer treatment, there is an increasing demand for accurate and comprehensive profile of tumor immunogenomics landscape in order to broaden our knowledge on predicting patient response as well as to facilitate drug development process. We established a cloud based analytical platform to comprehensively characterize genomic features that are potentially related to patients’ responses to immunotherapy. We implemented a neoantigen prediction pipeline based on whole exome sequencing (WES) and transcriptome sequencing data that integrates accurate HLA typing based on WES, somatic variant detection/phasing, RNA expression, class I MHC binding prediction as well as peptide epitope ranking. Among that, the HLA typing pipeline was validated using 73 Chinese and Japanese subjects covering >60 known class I HLA alleles with an accuracy above 95%. In addition, we also developed and validated methods to compute microsatellite instability (MSI) and tumor mutation burden (TMB) based on either a 600 gene panel or WES. Thirdly, this platform also enables the detection of HLA somatic variants and loss of heterozygosity events based on WES data. Next, we applied this analysis platform to process 127 samples across four cancer types (colon, rectal, endometrial and gastric) from TCGA as well as two Chinese cohorts of 100 non-small cell lung cancers and 30 esophageal squamous cell carcinomas sequenced at WuXi NextCODE. Within the TCGA cohort, the data reveals that patients with MSI High status produces significantly more neoantigens than patients with MS Stable status (7.06±4.60 vs 0.71±0.64, p = 7.65e-16, neo-peptides per Mb). Number of neopeptides predicted from each frameshift event is significantly higher than that from each missense mutation (1.30±1.56 vs 0.58±0.54, p = 2.86e-11, neo-peptides per Mb). We also observed strong correlation between TMB and neoantigen burden across multiple cancer types. In conclusion, we have developed a immunogenomics solution that captures tumors’ TMB, MSI, neoantigen as well as deleterious events associated with their HLAs. This platform will allow more comprehensive molecular features assessed to potentially improve the prediction of patients’ response to immune therapy.

Citation Format: Jie Wang, Ao Li, Jiawei Wang, Zhaoze Cheng. A comprehensive immunogenomics profiling platform enables the exploration of the intricate relationship between TMB, MSI, Neoantigen and HLA status among multiple cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 754.
Abstract 770: MiR628-5p targets Jagged 1 and inhibits growth, stemness and invasiveness of advanced prostate cancer cells
Authors: Rios-Colon Leslimar , Chijioke Juliet , Srivastava Anvesha , Ramalinga Malathi , Kedir Habib , Arthur Elena , Cagle Patrice , Deep Gagan , Kumar Deepak
Published: August 2019

Prostate cancer (PCa) is the most common non-cutaneous cancer in men, and according to the American Cancer Society reports, 164,690 new cases and 29,430 deaths from PCa are estimated in the United States in 2018. To reduce PCa mortality, we need a better understanding of molecular regulators of prostate carcinogenesis. MicroRNAs (MiRs) are small, non-coding RNA molecules that have been implicated in various cancers, and offer a novel opportunity to diagnose and treat cancer. We have previously identified MiR628-5p as a potential biomarker in PCa patients. This miRNA was found to be downregulated in the serum of both African American and Caucasian men with PCa compared to healthy individuals. In the present study, we further characterized the anti-cancer role of MiR628-5p in various PCa cell culture models. Results showed that MiR628-5p mimic treatment (100 nM) for 5 days significantly decreased the cell proliferation (~43% inhibition, p<0.001) in human prostate carcinoma PC3 cells compared to random mimic control. Similarly, MiR628-5p mimic treatment decreased the number of clones (clone size ≥50 cells) formed by PC3 cells by 40% (p<0.001) compared to random mimic control. Interestingly, miRNA 628-5p mimic treatment also reduced the stemness of PC3 cells as reflected in significant decrease in prostasphere size and CD44 expression. In mechanistic studies, we identified Jagged-1 as a molecular target regulated by MiR628-5p. MiR628-5p mimic treatment strongly decreased the Jagged-1 expression in several PCa cell lines (PC3, DU145, LNCaP and C4-2) as well as reduced the Jagged-1 luciferase activity. Jagged 1 inhibition by MiR628-5p was associated with reduced Notch signaling as well as decreased expression of transcription factors (snail and slug) regulating EMT. Moreover, MiR628-5p mimic treatment increased the expression of epithelial biomarkers (E-cadherin and ZO1), reduced the expression of mesenchymal marker (vimentin) and strongly inhibited the invasiveness of PC3 cells. In conclusion, we present evidence that MiR628-5p is a key regulator of prostate carcinogenesis, and offers a novel therapeutic opportunity to inhibit the growth of advanced PCa.

Citation Format: Leslimar Rios-Colon, Juliet Chijioke, Anvesha Srivastava, Malathi Ramalinga, Habib Kedir, Elena Arthur, Patrice Cagle, Gagan Deep, Deepak Kumar. MiR628-5p targets Jagged 1 and inhibits growth, stemness and invasiveness of advanced prostate cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 770.
Abstract 774: Regulation of autophagy and epigenetic modulator micro RNA by transient receptor potential melastatin 7 (TRPM7) channel in glioma
Authors: Liu Mingli
Published: August 2019

High grade malignant gliomas are devastating, uniformly fatal cancers for which no effective therapies currently exist. Glioblastoma multiforme (GBM) is the most common and aggressive primary brain tumor. We have reported that TRPM7 channels, a subfamily member of the transient receptor potential (TRP), regulate glioma stem cell (GSC) growth/proliferation through STAT3 and Notch signaling pathways. Autophagy is a catabolic process that occurs during both physiological and pathological process which has been reported to have dual roles, functioning either as a tumor-suppressor or tumor-promoter in cancer, including glioma. In addition, because of the limited success of the traditional treatment in prolonging the overall survival in GBM patients, epigenetics and epigenetic modulators have been attracting an increasing amount of attention for their influence in many aspects of cancer. microRNAs (miRNA), which can be considered as epigenetic effectors, reversibly regulate transcription through binding to complementary sequences of mRNA and silencing its translation into proteins. Therefore, we first determined the role of autophagy process in regulating glioma by TRPM7 using real time RT2 Profile PCR arrays (SABioscience, PAHS-084Z). We found that ATG16L1, ATG9A, CDKN1B, CLN3, CXCR4, and RGS19 were upregulated in response to TRPM7 silencing with the fold change greater than 1.6 using the A172 cell line of human glioma. The gene changes were further validated in other three glioma cell lines U87MG, U373MG and SNB19, which demonstrated different glioma tumorigenicity. miroRNA microarray analysis were then were performed upon TRPM7 silencing in A172 cells (www.LCsciences.com). In-depth data analysis from miRNA microarray data showed a list of 16 downregulated and 10 upregulated miRNA whose transcripts are statistically significant with fold change greater than 2 by TRPM7 knock-down. Among these, the micro RNA of hsa-miR-26b-5p, has-miR4530, and has-miR-28-5p have shown to exert crucial influence on tumor growth and migration by modulating AKT, ERK and IGF-1 signaling pathways. Stem-loop RT-qPCR were then used to detect and quantify miRNAs identified in miRNA microarray data from U87MG, U373MG and SNB19 cell lines. Our data indicate that autophagy effects serve as tumor suppressors in gliomagenesis and are reversely regulated by TRPM7; while hsa-miR-26b-5p, has-miR4530, and has-miR-28-5p are regulated by TRPM7 and participate in glioma progression.

Citation Format: Mingli Liu. Regulation of autophagy and epigenetic modulator micro RNA by transient receptor potential melastatin 7 (TRPM7) channel in glioma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 774.
Abstract 786: Metastasis is driven by mitochondrial protein A via mitochondrial reprogramming in colorectal cancer
Authors: Kim Dayeon , Kwon Sang-Mo
Published: August 2019

Colorectal cancer (CRC) is the third most common cancer worldwide, and CRC metastasis is the main cause of cancer-associated mortality. The hypoxia signaling pathway controls mediators to facilitate cancer progression and metastasis. Recently, mitochondrial reprogramming has emerged as a one of the mechanisms of hypoxia-induced metastasis; however, the link between hypoxia-induced metastasis and mitochondrial reprogramming remains to be elucidated. Here, we have focused on the novel mitochondrial protein A, to determine its role in the molecular mechanism of hypoxia-induced metastasis. Our results indicate that mitochondrial protein A is a downstream target of hypoxia-inducible factor 1 (HIF-1α) and plays an essential role in HIF-1-induced migration and invasion. In addition, we found a clinical correlation between hypoxia and mitochondrial protein A. To identify the role of mitochondrial protein A, we generated mitochondrial protein A deficient or silenced cells, which showed impaired migration, invasion, and colony-forming ability even under hypoxic condition. In contrast, ectopic expression of mitochondrial protein A elevated metastatic potential, both in vitro and in vivo. Mechanistically, mitochondrial protein A directly interacted with dynamin-related protein (DRP1) to retain it in mitochondria, thereby promoting mitochondrial fission. To confirm the effect of mitochondrial fission on mitochondrial protein A-induced metastasis, we blocked DRP1 in mitochondrial protein A overexpressing cells by pharmacological inhibition, which led to impairment of mitochondrial protein A-induced metastatic potential. Furthermore, mitochondrial protein A-silenced cells showed decreased metabolism and lactate production, suggesting that mitochondrial protein A regulates the mitochondrial reprogramming. In summary, we demonstrate that mitochondrial protein A is a novel DRP1 adaptor protein, facilitating mitochondrial fission and mitochondrial reprogramming, and plays a critical role in hypoxia-induced metastasis. We suggest that targeting of mitochondrial protein A maybe a strategy for cancer therapy.

Citation Format: Dayeon Kim, Sang-Mo Kwon. Metastasis is driven by mitochondrial protein A via mitochondrial reprogramming in colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 786.
Abstract 794: Inhibition of mitochondrial metabolism by a novel agent targeting RUNX2-DNA binding in breast cancer
Authors: Kim Myoung Sook , Gernapudi Ramkishore , Cedeno Yessenia C. , Polster Brian M. , Girnun Geoffrey D. , Passaniti Antonino
Published: August 2019

The RUNX2 transcription factor promotes breast cancer growth and metastasis through interactions with a variety of cofactors that activate or repress target genes. RUNX2 knockdown decreases expression levels of glycolytic genes such as LDHA, HK2 and GLUT-1, but upregulates PDHA1 mRNA level and enzymatic activity, which is consistent with lower glycolytic potential. Using a direct computer-assisted drug discovery (CADD) approach, we identified CADD522 as a small molecule that inhibits the RUNX2-DNA binding, transcriptional activity of RUNX2, and expression of RUNX2 target genes. We also observed that CADD522 inhibits breast cancer growth and metastasis using in vitro and in vivo tumor models. RUNX2-activated glucose utilization and GLUT-1 levels were also reduced by CADD522. New data also define CADD522 as a potential antitumor agent that downregulates the levels of metabolites in the TCA cycle and inhibits mitochondrial oxygen consumption rates. Our results support the observations that CADD522 could regulate cellular metabolism through modulation of both glycolytic metabolism and mitochondrial oxidative phosphorylation. To address the mechanisms by which CADD522 exerts its inhibitory effects on mitochondrial metabolism, we performed RNA-Sequencing analysis in breast cancer cells and identified several specific molecular signatures that determine cellular response to RUNX2 and CADD522. Further pathway changes in mitochondrial function as well as validation of differentially expressed genes that play critical roles in mitochondrial metabolism are under investigation.

Citation Format: Myoung Sook Kim, Ramkishore Gernapudi, Yessenia C. Cedeno, Brian M. Polster, Geoffrey D. Girnun, Antonino Passaniti. Inhibition of mitochondrial metabolism by a novel agent targeting RUNX2-DNA binding in breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 794.
Abstract 797: Obesity-associated, but not obesity-independent, tumors respond to insulin by increasing mitochondrial glucose oxidation
Authors: Rabin-Court Aviva , Shulman Gerald I. , Perry Rachel J.
Published: August 2019

Obesity is associated with increased incidence and worse prognosis of more than one dozen tumor types; however, the molecular mechanisms for this association are unknown. We hypothesized that insulin, which is elevated in obesity-driven insulin resistance, would increase tumor glucose oxidation in obesity-associated tumors. To test this hypothesis, we developed a stable isotope method in which tumor cells are incubated in physiological concentrations (5 mM) of [13C6] glucose and the ratio of pyruvate dehydrogenase flux to citrate synthase flux (VPDH/VCS, i.e. the percent of total mitochondrial oxidation fueled by glucose oxidation) is given as [4,5-13C2]glutamate/[13C3]alanine, measured by gas chromatography/mass spectrometry and liquid chromatography-tandem mass spectrometry respectively. Using this method, we found that both breast cancer (4T1) and colon cancer (MC38) cells, tumors that are associated with obesity, exhibit greater relative rates of glucose oxidation to total mitochondrial oxidation (VPDH/VCS 61±3% and 60±2%, respectively) than melanoma (YUMM) and Renca (renal cell carcinoma) cells (41±4% and 37±2%, respectively; P<0.05 for all comparisons between obesity-associated and non-obesity-associated tumors). In addition, both 4T1 and MC38 cells increased VPDH/VCS upon incubation in 100 nM insulin (75±3% and 81±2%, respectively, P<0.01 vs. the same tumor type without insulin), whereas there was no change in glucose oxidation with insulin in YUMM or Renca cells (35±3% and 36±2%, respectively). These data reveal that a shift in substrate preference may comprise a metabolic signature of obesity-associated tumors that differs from that of those not associated with obesity. In addition, contrary to the conventionally held view that glucose oxidation in tumor cells is constitutively high, these data reveal that obesity-driven tumors respond acutely to insulin by increasing mitochondrial glucose oxidation, whereas obesity-independent tumor cells do not. CONCLUSION: These data demonstrate that obesity associated tumors (breast and colon), in contrast to tumors that are not associated with obesity (melanoma, renal cell), are insulin-responsive and have higher rates of basal and insulin-stimulated mitochondrial glucose oxidation. Therefore assessment of in vitro tumor mitochondrial substrate preference may predict whether interventions to reverse hyperinsulinemia or reduce insulin-responsive glucose oxidation in tumors may be attractive therapeutic modalities in cancer types associated with obesity.

Citation Format: Aviva Rabin-Court, Gerald I. Shulman, Rachel J. Perry. Obesity-associated, but not obesity-independent, tumors respond to insulin by increasing mitochondrial glucose oxidation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 797.
Abstract 80: Electronic cigarette aerosols increase clonogenic survival in oral cancer cells after cisplatin treatment
Authors: Manyanga Jimmy , Mehta Toral R. , Queimado Lurdes
Published: August 2019

Background and Aim: Are electronic cigarettes (ECs) safer than combustible tobacco? No one knows the answer yet. ECs are devices that deliver an aerosol from a heated mixture of propylene glycol, vegetable glycerin, and flavors, with or without nicotine. Short-term EC studies are mostly limited and inconclusive. Despite unknown long-term health effects, the use of ECs has increased alarmingly over the past decade. EC aerosols contain unique chemicals as well as other chemicals similar to those found in tobacco smoke. Tobacco smoking during cancer treatment has been associated with increased drug resistance and reduced overall survival rate. The awareness of the dangers of cigarette smoking and the paucity of studies evaluating the safety of ECs pose a dilemma for cancer patients who might be considering switching to ECs during chemotherapy. Here, we sought to examine the effects of EC aerosol extracts exposure on cell survival in oral cancer cells during cisplatin treatment.

Methods: EC aerosol extracts were collected from two brands of ECs as previously described. Mainstream tobacco smoke (MS) extract was used as a positive control. To evaluate clonogenic survival after cisplatin treatment, oral cancer cells were exposed for 48 h to EC aerosol extracts at nicotine doses comparable to those observed in EC users. Next, cells were treated with both EC aerosol extracts and cisplatin (or vehicle-control) for another 48 h. After treatment, cells were counted and seeded in 6-well plates and colony formation was assessed two weeks later. Colonies were fixed with methanol followed by staining with 0.5% crystal violet in 25% methanol and counted manually. Only colonies with at least 50 cells were counted. Data were analyzed by Student's t-tests and one-way analysis of variance (ANOVA).

Results: After cisplatin treatment, cells exposed to EC aerosol extracts showed a significant increase in clonogenic survival fraction compared to vehicle-control treated cells. Furthermore, cells exposed to EC aerosol extracts formed visibly larger colonies. An increase in clonogenic survival was also observed in MS extract-treated cells.

Conclusions: Collectively, our data suggest that short-term exposure to EC aerosol can induce cisplatin resistance in oral cancer cells, thus allowing the cells to survive and form colonies. These findings are particularly important in paving the way to understanding whether ECs are a safer alternative for oral cancer patients undergoing chemotherapy.

Citation Format: Jimmy Manyanga, Toral R. Mehta, Lurdes Queimado. Electronic cigarette aerosols increase clonogenic survival in oral cancer cells after cisplatin treatment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 80.
Abstract 802: Effective breast cancer combination therapy targeting BACH1 and mitochondrial metabolism
Authors: Lee Jiyoung
Published: August 2019

Mitochondrial metabolism is an attractive target for cancer therapy. Reprogramming metabolic pathways could improve the ability of metabolic inhibitors to suppress cancers with limited treatment options like triple negative breast cancer (TNBC). Here we show that BACH1 (BTB and CNC homology1), a heme-binding transcription factor whose expression is enriched in TNBC patients, targets mitochondrial metabolism. BACH1 decreases glucose utilization in the TCA cycle and negatively regulates transcription of electron transport chain (ETC) genes. BACH1 depletion by shRNA or degradation by hemin sensitizes cells to ETC inhibitors such as metformin, suppressing growth of both cell line and patient-derived tumor xenografts. Expression of a heme-resistant BACH1 mutant in shBACH1 cells rescues the BACH1 phenotype and restores metformin resistance in hemin-treated cells and tumors. Finally, BACH1 gene expression inversely correlates with ETC gene expression in breast cancer patients as well as other tumor types, highlighting the clinical relevance. This study demonstrates that mitochondrial metabolism can be exploited through targeting BACH1 to sensitize breast cancer and potentially other tumor tissues to mitochondrial inhibitors.

Citation Format: Jiyoung Lee. Effective breast cancer combination therapy targeting BACH1 and mitochondrial metabolism [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 802.
Abstract 803: Identification of compensatory pathway for glutamate production
Authors: Asaka Ryoichi , Nguyen Tu , Krisch Brian J. , Nabi Karim , Quinones Addison , Tan Jessica , Antonio Marjorie J. , Li Ting , Camelo Felipe , Nguyen Kiet , Udupa Sunag , Gabrielson Edward , Le Anne
Published: August 2019

Measurement and Methods: In this study, we sought to expand our knowledge of glutamine metabolism beyond glutaminolysis. Using mass spectroscopy-based stable isotope-resolved metabolomics (SIRM) with 13C515N2-labeled-glutamine, we precisely identified the metabolites produced from glutamine both in vitro and in vivo. Tumors were harvested 2 hours post first glutamine injection. Metabolites were then extracted from tumors, and analyzed using Agilent 6520 Q-TOF mass spectrometer and 1H-NMR. Metabolite intensities were later normalized to protein concentration following analysis.

Results: Our results showed that total glutamate levels were lower in BPTES-NP treated tumors as compared to vehicle control ones. Interestingly, we found an increase in (m+5) labeled glutamate (mass of the parent ions (m) and 5 more mass units due to 13C415N1-glutamate or 13C5-glutamate labelling) in BPTES-NP treated tumors as compared to the vehicle control tumors. Moreover, we found that (m+5) glutamate is a product of (m+7) glutamine being converted to (m+5) alpha-ketoglutaramate (KGM) via glutamine-pyruvate transaminase and further on into alpha-ketoglutarate (aKG) by omega-amidase, which can finally produce the identified (m+5) glutamate through glutamate dehydrogenase. Further analysis using 1H-NMR detailed a significant increase in overall KGM intermediate intensity in treatment groups compared to the control groups, confirming the upregulation of compensatory pathway (glutamine-KGM- aKG-glutamate) to produce glutamate upon treatment of GLS1 inhibitor.

Conclusion: These results explain the reasons behind the limited clinical outcomes for single therapy with a GLS1 inhibitor, and provide potential therapeutic targets: glutamine-pyruvate transaminase, for combination treatments with GLS1 inhibitors to prevent the compensation for glutamate production amid GLS1 inhibition.

Citation Format: Ryoichi Asaka, Tu Nguyen, Brian J. Krisch, Karim Nabi, Addison Quinones, Jessica Tan, Marjorie J. Antonio, Ting Li, Felipe Camelo, Kiet Nguyen, Sunag Udupa, Edward Gabrielson, Anne Le. Identification of compensatory pathway for glutamate production [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 803.
Abstract 810: Differential use of lactate for mitochondria respiration by NSCLC cells
Authors: Swain Pamela , Romero Natalia , Kam Yoonseok , Dranka Brian P.
Published: August 2019

Metabolic liabilities in cancer cells provide the opportunity for novel therapeutic approaches. This is especially true as the dogma that all cancer cells are glycolytic is being challenged. Phenotypic screening of cancer cell lines can demonstrate what these liabilities are and suggest approaches for drugs that target these vulnerabilities. As an example, non-small cell lung cancer (NSCLC) is typically driven by oncogenic mutations in either KRAS or EGFR. However, the impact of these mutations on cellular metabolic phenotype is not well-studied. In this study, we observed that the two KRAS-mutated NSCLC cell lines (A549 and H460) primarily relied on glycolysis for ATP production while EGFR-mutated cell lines (H1975 and PC9) were more reliant on mitochondrial respiration to meet energy demands. There is emerging evidence to suggest that lactate is a major fuel for cancer cell energy metabolism especially in the glucose-limited tumor microenvironment. Thus, we next compared the use of lactate with other carbon sources for ATP production in NSCLC cells. We found there was differential usage of lactate for mitochondrial respiration between KRAS and EGFR-mutated NSCLC cells. EGFR-mutated NSCLC cells used lactate and increased mitochondrial respiration when lactate was acutely administrated regardless of glucose availability. In contrast, the use of lactate for the mitochondrial respiration by KRAS-mutated cells was limited. In A549 cells, no significant change in mitochondrial respiration rate was observed at all by acute injection of lactate. Together, these results imply that NSCLC variants have adopted different metabolic phenotypes depending on the oncogenic background and more oxidative phenotype may be correlated to higher lactate use for TCA cycle in production of ATP.

Citation Format: Pamela Swain, Natalia Romero, Yoonseok Kam, Brian P. Dranka. Differential use of lactate for mitochondria respiration by NSCLC cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 810.