Rapid Tumor Cell Proliferation Research Articles

Research Advances in Antitumor Mechanism of Evodiamine

Evodiamine is a natural alkaloid extracted from Fructus Evodia. This bioactive alkaloid has been reported to have a wide range of biological activities, including anti-injury, antiobesity, vasodilator, and anti-inflammatory effects. In recent years, it has been found that evodiamine has tumor-suppressive effects on a variety of tumors. There is growing evidence that evodiamine can inhibit the rapid proliferation of tumor cells, induce cell cycle arrest at a certain phase, increase the incidence of apoptosis, promote autophagy, inhibit microangiogenesis and migration, and regulate immunotherapy. Evodiamine can inhibit Wnt/β-catenin, mTOR, NF-κB, PI3K/AKT, JAK-STAT, and other signaling pathways in various cancer cells, and it can significantly downregulate the expression of many tumor markers, such as VEGF and COX-2. These facts partially explain the antitumor mechanism of evodiamine. In this article, the antitumor mechanism of evodiamine was reviewed to provide the basis for its clinical application and therapeutic development in the future.

Metabolomic Phenotyping of Gliomas: What Can We Get with Simplified Protocol for Intact Tissue Analysis?

Simple SummaryThe diagnostic protocol for gliomas is based on histological examination and the determination of genetic biomarkers. However, examining molecular biomarkers in cancer tissue is usually labor-intensive and time-consuming when a homogenization step is involved. Therefore, this diagnostic approach has not been fully explored to date. The present study seeks to validate the applicability of solid-phase microextraction (SPME), or chemical biopsy, as a new approach for fast and simple sampling and sample-preparation in the surgery room prior to the application of metabolomic analysis to identify biomarkers. To this end, the metabolomic profiles of brain tumors were compared with genetic biomarkers and the results of histological analysis in order to identify changes of molecular metabolites of statistical significance. The findings of this study indicate that the proposed approach provides complementary information to current diagnostic methods and has the potential to be a valuable on-site analytical tool in future applications.Glioblastoma multiforme is one of the most malignant neoplasms among humans in their third and fourth decades of life, which is evidenced by short patient survival times and rapid tumor-cell proliferation after radiation and chemotherapy. At present, the diagnosis of gliomas and decisions related to therapeutic strategies are based on genetic testing and histological analysis of the tumor, with molecular biomarkers still being sought to complement the diagnostic panel. This work aims to enable the metabolomic characterization of cancer tissue and the discovery of potential biomarkers via high-resolution mass spectrometry coupled to liquid chromatography and a solvent-free sampling protocol that uses a microprobe to extract metabolites directly from intact tumors. The metabolomic analyses were performed independently from genetic and histological testing and at a later time. Despite the small cohort analyzed in this study, the results indicated that the proposed method is able to identify metabolites associated with different malignancy grades of glioma, as well as IDH and 1p19q codeletion mutations. A comparison of the constellation of identified metabolites and the results of standard tests indicated the validity of using the characterization of one comprehensive tumor phenotype as a reflection of all diagnostically meaningful information. Due to its simplicity, the proposed analytical approach was verified as being compatible with a surgical environment and applicable for large-scale studies.

Oxygen nanocarrier broke the hypoxia trap of solid tumors and rescued transfection efficiency for gene therapy

BackgroundGene therapy shows great promise for a broad array of diseases. However, we found that hypoxic tumor microenvironment (TME) exerted significant inhibitory effects on transfection efficiency of a variety of gene vectors (such as Lipo 2000 and PEI) in an oxygen-dependent manner. Solid tumors inevitably resulted in acute hypoxic areas due to the rapid proliferation of tumor cells and the aberrant structure of blood vessels. Thus, the hypoxic TME severely limited the efficiency and application of gene therapy.MethodsIn our previous study, we constructed endoplasmic reticulum-targeted cationic liposomes, PAR-Lipo, which could effectively deliver genes and ensure high transfection efficiency under normoxia. Unsatisfactorily, the transfection efficiency of PAR-Lipo was rather poor under hypoxia. We believed that reoxygenation was the most direct and effective means to rescue the low transfection under hypoxia. Hence, we fabricated liposomes modified with perfluorooctyl bromide (PFOB@Lipo) to load oxygen and deliver it to tumor sites, which effectively alleviated the hypoxic nature of tumor. Then PAR-Lipo were applied to mediate high-efficiency delivery of tumor suppressor gene pTP53 to inhibit tumor progression.ResultsThe results showed that such staged strategy augmented the expression of P53 protein in tumors and extremely suppressed tumor growth.ConclusionThis work was the first attempt to utilize an oxygen nanocarrier to assist the therapeutic effect of gene therapy under hypoxia, providing a new reference for gene therapy in malignant tumors.Graphical Abstarct

MYCN recruits the nuclear exosome complex to RNA polymerase II to prevent transcription-replication conflicts

The MYCN oncoprotein drives the development of numerous neuroendocrine and pediatric tumors. Here we show that MYCN interacts with the nuclear RNA exosome, a 3'-5' exoribonuclease complex, and recruits the exosome to its target genes. In the absence of the exosome, MYCN-directed elongation by RNA polymerase II (RNAPII) is slow and non-productive on a large group of cell-cycle-regulated genes. During the S phase of MYCN-driven tumor cells, the exosome is required to prevent the accumulation of stalled replication forks and of double-strand breaks close to the transcription start sites. Upon depletion of the exosome, activation of ATM causes recruitment of BRCA1, which stabilizes nuclear mRNA decapping complexes, leading to MYCN-dependent transcription termination. Disruption of mRNA decapping in turn activates ATR, indicating transcription-replication conflicts. We propose that exosome recruitment by MYCN maintains productive transcription elongation during S phase and prevents transcription-replication conflicts to maintain the rapid proliferation of neuroendocrine tumor cells.

Emerging Roles of Energy Metabolism in Ferroptosis Regulation of Tumor Cells.

Ferroptosis is a new form of regulated cell death, which is characterized by the iron‐dependent accumulation of lethal lipid peroxides and involved in many critical diseases. Recent reports revealed that cellular energy metabolism activities such as glycolysis, pentose phosphate pathway (PPP), and tricarboxylic acid cycle are involved in the regulation of key ferroptosis markers such as reduced nicotinamide adenine dinucleotide phosphate (NADPH), glutathione (GSH), and reactive oxygen species (ROS), therefore imposing potential regulatory roles in ferroptosis. Remarkably, tumor cells can activate adaptive metabolic responses to inhibit ferroptosis for self‐preservation such as the upregulation of glycolysis and PPP. Due to the rapid proliferation of tumor cells and the intensified metabolic rate, tumor energy metabolism has become a target for disrupting the redox homeostasis and induce ferroptosis. Based on these emerging insights, regulatory impact of those‐tumor specific metabolic aberrations is systematically characterized, such as rewired glucose metabolism and metabolic compensation through glutamine utilization on ferroptosis and analyzed the underlying molecular mechanisms. Additionally, those ferroptosis‐based therapeutic strategies are also discussed by exploiting those metabolic vulnerabilities, which may open up new avenues for tumor treatment in a clinical context.

Clinical Significance and Immunologic Landscape of a Five-IL(R)-Based Signature in Lung Adenocarcinoma.

Interleukins (ILs) and interleukin receptors (ILRs) play important role in the antitumor immune response. However, the expression signature and clinical characteristics of the IL(R) family in lung adenocarcinoma (LUAD) remains unclear. The main purpose of this study was to explore the expression profile of IL(R) family genes and construct an IL(R)-based prognostic signature in LUAD. Five public datasets of 1,312 patients with LUAD were enrolled in this study. Samples from The Cancer Genome Atlas (TCGA) were used as the training set, and samples from the other four cohorts extracted from Gene Expression Omnibus (GEO) database were used as the validation set. Additionally, the profile of IL(R) family signature was explored, and the association between this signature and immunotherapy response was also analyzed. Meanwhile, the prognostic value was compared between this IL(R)-based signature and different immunotherapy markers. A signature based on five identified IL(R)s (IL7R, IL5RA, IL20RB, IL11, IL22RA1) was constructed using the TCGA dataset through univariate/multivariable Cox proportional hazards regression and least absolute shrinkage and selection operator (LASSO) Cox analysis. These cases with LUAD were stratified into high- and low-risk group according to the risk score. This signature showed a strong prognostic ability, which was verified by the five independent cohorts and clinical subtypes. The IL(R)-based models presented unique characteristics in terms of immune cell infiltration and immune inflammation profile in tumor microenvironment (TME). Biological pathway analysis confirmed that high-risk patients showed significant T- and B-cell immunosuppression and rapid tumor cell proliferation. More importantly, we researched the relationship between this IL(R)-based signature and immune checkpoints, tumor mutation burden (TMB), tumor purity and ploidy, and tumor immune dysfunction and exclusion (TIDE) score, which confirmed that this signature gave the best prognostic value. We first provided a robust prognostic IL(R)-based signature, which had the potential as a predictor for immunotherapy response to realize individualized treatment of LUAD.

Abstract 2322: Targeting glutamine transporter (ASCT2) inhibits metabolic stress-induced GD2+ cancer stem cell-like phenotype in triple-negative breast cancer

Abstract Nutrient deprivation is one of the hallmark conditions of a rapidly growing tumor. Rapid proliferation of tumor cells leads to increased consumption of metabolites such as glucose and glutamine resulting in metabolic stress. However, phenotypic and functional status of tumor cells under these conditions is not clear. We hypothesize that nutrient deprivation-induced metabolic stress results in a breast cancer stem cell-like (BCSC) phenotype in triple negative breast cancer (TNBC). We found that serum-deprived conditions result in significant upregulation of GD2+ cells compared to serum rich conditions: from 9%±2% to 26%±5% in MDA-MB-231 and 7%±2% to 29%±5% in SUM159 cells. Interestingly, this affect was compensated when the cells were supplemented with serum-rich medium. In addition, a positive correlation was observed between GD2+ cell number and tumor volume in vivo, suggesting metabolic stress in the growing tumors results in GD2+ phenotype in TNBC cells. Next, TNBC cells cultured under glucose deprivation or in the presence of glycolysis inhibitor, 2-deoxy glucose (2DG) showed a 2-fold increase in GD2+ cells compared to cells cultured with glucose (6g/L) or without 2DG, suggesting that glucose deprivation enhances GD2 expression. Nutrient deprived conditions also showed a significant increase in the expression of stress markers ATF4 and SESN2 demonstrating TNBC cells under metabolic stress. In addition, the above conditions also showed a 2.5-fold upregulation of the mammosphere formation in TNBC cells compared to nutrient rich conditions. Global metabolomics profiling identified glutathione biosynthesis to be most highly upregulated in GD2+ cells. Furthermore, we found that supplementation of glutamine (precursor for glutathione biosynthesis) increases the percentage of GD2+ cells. Carbon and nitrogen tracing using 13C- and 15N-labeled glutamine by mass spectrometry showed its direct contribution to GD2 biosynthesis. Treatment with glutamine transporter (ASCT2) inhibitor V9302 reduced GD2+ cell number by 70%-80% in TNBC cell lines and dramatically inhibited mammosphere and soft agar colony formation, and migration of TNBC cell lines. Mechanistic studies revealed that V9302 inhibits glutathione pathway and induces ROS levels in TNBC cells. Additionally, V9302 treatment inhibited AKT-ERK-mTOR-4E-BP1 pathway and combination of V9302 with paclitaxel synergizes TNBC cell killing. We also identified that, V9302 treatment induced ferroptosis (iron-dependent cell death) in TNBC cells. Finally, V9302 treatment significantly inhibited tumor growth (p<0.0001) in TNBC-PDX model. In conclusion, metabolic stress results in GD2+ BCSC phenotype in TNBC cells and glutamine directly contributes to GD2 biosynthesis. Targeting ASCT2 could complement conventional chemotherapy in TNBC. Citation Format: Appalaraju Jaggupilli, Stanley Ly, Khoa Nguyen, Roshan Borkar, Bin Yuan, Danthasinghe Waduge Piyarathna, Yuanqing Yan, Helen Piwnica-Worms, Henry Charles Manning, Michael Andreeff, Nagireddy Putluri, Lokesh Battula. Targeting glutamine transporter (ASCT2) inhibits metabolic stress-induced GD2+ cancer stem cell-like phenotype in triple-negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2322.

Selective Anti-melanoma Effect of Phosphothioated Aptamer Encapsulated by Neutral Cytidinyl/Cationic Lipids.

BC15-31 is a DNA aptamer that targets heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1), which plays a crucial role in the process of pre-RNA maturation and is also essential for the rapid proliferation of tumor cells. In this research, we modified BC15-31 with a phosphorothioate (PS) backbone, LNA, and 2-O-MOE to enhance its stability and target affinity. In addition, a neutral cytidinyl lipid (DNCA) and a cationic lipid (CLD) were mixed to encapsulate modified aptamers with the aim of improving their cell permeability with low toxicity. Under the DNCA/CLD package, aptamers are mainly distributed in the nucleus. A modified sequence WW-24 showed an excellent selective anti-melanoma (A375 cells, ∼25 nM, 80%) activity, targeted to both hnRNP A1 and hnRNP A2/B1 found by the BLI experiment, and induced more early and late apoptosis in vitro, which also showed stronger antitumor effect and longer accumulation time in vivo. These results provide a new strategy for further clinical applications.

Photo-responsive hydrogel facilitates nutrition deprivation by an ambidextrous approach for preventing cancer recurrence and metastasis

Postoperative recurrence at the primary site and distant metastasis remains the challenge in treating triple-negative breast cancer due to its unpredictable invasion into adjacent tissues. Although systemic chemotherapy has been extensively adopted to attenuate the recurrence and metastasis, the abundant nutrition supply by blood vessels would promote the rapid proliferation of tumor cells and angiogenesis. Herein, we reported a nutrition deprivation strategy by ambidextrously blocking the residual blood vessels and inhibiting angiogenesis to realize efficient treatment of triple-negative breast cancer. To this end, an injectable hydrogel with photo-responsive property was prepared by using polydopamine crosslinked collagen/silk fibroin composite to deliver thrombin for blocking blood vessels and angiogenesis. In the presence of NIR light, the locked thrombin was released into the blood vessels in the adjacent tissues to promote blood coagulation. In addition, the photothermal effect would reduce the secreting of VEGF for preventing angiogenesis in the adjacent tissues. The in vitro and in vivo results demonstrated that the permanent interruption of nutrient supply by blocking the blood vessels adjacent to the resected tumor and preventing angiogenesis is a promising strategy to prevent the recurrence and metastasis of TNBC.

Prohibitin regulates mTOR pathway via interaction with FKBP8.

The ability of tumor cells to sustain continuous proliferation is one of the major characteristics of cancer. The activation of oncogenes and the mutation or inactivation of tumor suppressor genes ensure the rapid proliferation of tumor cells. The PI3K-Akt-mTOR axis is one of the most frequently modified signaling pathways whose activation sustains cancer growth. Unsurprisingly, it is also one of the most commonly attempted targets for cancer therapy. FK506 binding protein 8 (FKBP8) is an intrinsic inhibitor of mTOR kinase that also exerts an anti-apoptotic function. We aimed to explain these contradictory aspects of FKBP8 in cancer by identifying a "switch" type regulator. We identified through immunoprecipitation-mass spectrometry-based proteomic analysis that the mitochondrial protein prohibitin 1 (PHB1) specifically interacts with FKBP8. Furthermore, the downregulation of PHB1 inhibited the proliferation of ovarian cancer cells and the mTOR signaling pathway, whereas the FKBP8 level in the mitochondria was substantially reduced. Moreover, concomitant with these changes, the interaction between FKBP8 and mTOR substantially increased in the absence of PHB1. Collectively, our finding highlights PHB1 as a potential regulator of FKBP8 because of its subcellular localization and mTOR regulating role.

Centrosomal Localization of RXRα Promotes PLK1 Activation and Mitotic Progression and Constitutes a Tumor Vulnerability.

Retinoid X receptor alpha (RXRα), a nuclear receptor of transcription factor, controls various physiological and pathological pathways including cellular growth, proliferation, differentiation, and apoptosis. Here, we report that RXRα is phosphorylated at its N-terminal A/B domain by cyclin-dependent kinase 1 (Cdk1) at the onset of mitosis, triggering its translocation to the centrosome, where phosphorylated-RXRα (p-RXRα) interacts with polo-like kinase 1 (PLK1) through its N-terminal A/B domain by a unique mechanism. The interaction promotes PLK1 activation, centrosome maturation, and mitotic progression. Levels of p-RXRα are abnormally elevated in cancer cell lines, during carcinogenesis in animals, and in clinical tumor tissues. An RXRα ligand XS060, which specifically inhibits p-RXRα/PLK1 interaction but not RXRα heterodimerization, promotes mitotic arrest and catastrophe in a tumor-specific manner. These findings unravel a transcription-independent action of RXRα at the centrosome during mitosis and identify p-RXRα as a tumor-specific vulnerability for developing mitotic drugs with improved therapeutic index.

647P - Selective Wnt/β-catenin small-molecule inhibitor CWP232228 impairs tumour growth of colon cancer

BackgroundAberrant activation of Wnt/β-catenin signaling is commonly found in colorectal cancers (CRCs) and is the prime event involved in the rapid proliferation of tumor cells in patients with advanced CRC. However, the limited effects of current therapeutics are unsatisfying, which highlights the importance of novel interventions in CRC chemo-treatment. We designed a highly potent small-molecule inhibitor CWP232228, which antagonizes binding of β-catenin to T cell factor in the nucleus. Thus, the present study aimed to explore the possibility of a selective small molecule β-catenin inhibitor, CWP232228, as a potential therapeutic drug in the treatment of CRCs. MethodsWe observed that CWP232228 at a concentration of as low as 1.0μm induced significant cytotoxic effectsand apoptosis in HCT116. Immunoblot analysis showed that CWP232228 treatment increased the release of cytochrome C, which led to the activation of apical (caspase-9) and effector caspases (caspase-7 and -3) resulting in the increased cleavage of PARP. ResultsIn addition, we observed that CWP232228 caused cell cycle arrest via attenuating the expressions of cyclin D2, D3, and survivin. Thus, we validated decreased expressions of Wnt/β-catenin targeted cell cycle progression regulators, aurora kinase A, c-Myc, cyclin D1, and microphthalmia-associated transcription factor. ConclusionsWe confirmed that CWP232228 targets β-catenin via decreased promoter activities, nuclear expressions of β-catenin. Lastly, we found CWP232228 also inhibited growth of xenografted colon cancer cells in NOD-scid IL2Rgammanull mice. Collectively, results in the current study showed a possibility for CWP232228 as a potential therapeutic drug in CRCs. Legal entity responsible for the studyThe authors. FundingHas not received any funding. DisclosureAll authors have declared no conflicts of interest.

Tumor Dormancy and Interplay with Hypoxic Tumor Microenvironment.

The tumor microenvironment is a key factor in disease progression, local resistance, immune-escaping, and metastasis. The rapid proliferation of tumor cells and the aberrant structure of the blood vessels within tumors result in a marked heterogeneity in the perfusion of the tumor tissue with regions of hypoxia. Although most of the tumor cells die in these hypoxic conditions, a part of them can adapt and survive for many days or months in a dormant state. Dormant tumor cells are characterized by cell cycle arrest in G0/G1 phase as well as a low metabolism, and are refractive to common chemotherapy, giving rise to metastasis. Despite these features, the cells retain their ability to proliferate when conditions improve. An understanding of the regulatory machinery of tumor dormancy is essential for identifying early cancer biomarkers and could provide a rationale for the development of novel agents to target dormant tumor cell populations. In this review, we examine the current knowledge of the mechanisms allowing tumor dormancy and discuss the crucial role of the hypoxic microenvironment in this process.

Targeting translation initiation by synthetic rocaglates for treating MYC-driven lymphomas.

MYC-driven lymphomas, especially those with concurrent MYC and BCL2 dysregulation, are currently a challenge in clinical practice due to rapid disease progression, resistance to standard chemotherapy and high risk of refractory disease. MYC plays a central role by coordinating hyperactive protein synthesis with upregulated transcription in order to support rapid proliferation of tumor cells. Translation initiation inhibitor rocaglates have been identified as the most potent drugs in MYC-driven lymphomas as they efficiently inhibit MYC expression and tumor cell viability. We found that this class of compounds can overcome eIF4A abundance by stabilizing target mRNA-eIF4A interaction that directly prevents translation. Proteome-wide quantification demonstrated selective repression of multiple critical oncoproteins in addition to MYC in B cell lymphoma including NEK2, MCL1, AURKA, PLK1, and several transcription factors that are generally considered undruggable. Finally, (−)-SDS-1–021, the most promising synthetic rocaglate, was confirmed to be highly potent as a single agent, and displayed significant synergy with the BCL2 inhibitor ABT199 in inhibiting tumor growth and survival in primary lymphoma cells in vitro and in patient-derived xenograft mouse models. Overall, our findings support the strategy of using rocaglates to target oncoprotein synthesis in MYC-driven lymphomas.

Identification and Verification of Two Novel Differentially Expressed Proteins from Non-neoplastic Mucosa and Colorectal Carcinoma Via iTRAQ Combined with Liquid Chromatography-Mass Spectrometry.

Recurrence or metastasis of colorectal cancer (CRC) is common following surgery and/or adjuvant therapy, particularly in patients with an advanced stage of the cancer. Identifying key molecular markers of CRC is beneficial for early diagnosis and early treatment, which may eventually improve the prognosis of patients with CRC. Isobaric mass tags for relative and absolute quantification (iTRAQ) in combination with multidimensional liquid chromatography and tandem mass spectrometry (LC-MS/MS) were used to identify differentially expressed proteins between CRC tissues and paired adjacent normal mucosa. Among the 105 patients, adenocarcinoma was the most common CRC subtype, stage III was the most common Tumor-Node-Metastasis stage and high levels of Ki-67 indicated the rapid proliferation of tumor cells in the samples. The LC-MS/MS-based iTRAQ technology identified 271 differentially expressed proteins, with 130 upregulated proteins and 141 downregulated proteins. Bioinformatics analysis revealed that golgin subfamily A member 2 (GOLGA2) and heterogeneous nuclear ribonucleoprotein D0 (hnRNPD) were located in the center of the upregulated protein network, and were closely associated with the development of CRC. The upregulation of GOLGA2 and hnRNPD was further verified in human tissues using western blotting and immunohistochemistry. GOLGA2 and hnRNPD were identified as two novels differentially expressed proteins in human CRC. Furthermore, the LC-MS/MS-based iTRAQ proteomic approach is a useful tool for searching and identifying differentially expressed proteins, and may be used to provide a comprehensive understanding of the processes that mediate the development of CRC.

TMIC-19. USING QUANTITATIVE MR IMAGING TO RELATE GBM MASS EFFECT TO PERFUSION AND DIFFUSION CHARACTERISTICS OF THE TUMOR MICRO-ENVIRONMENT

Biomechanical forces are known to affect tumor growth and evolution [1]. Likewise, tumor growth drives physical changes in the micro-environment that affect tissue solid and fluid mechanics. Tumor mass effect, resulting from rapid tumor cell proliferation, has been shown to be prognostic for poor outcome in glioblastoma (GBM) patients and to be associated with the expression of gene signatures consistent with proliferative growth phenotype [2]. Similarly, elevated interstitial fluid flow (IFF) has been shown to drive GBM invasion [3]. This study investigates the relationship between tumor mass effect, diffusion, perfusion and IFF in GBM using anatomical (pre- and post-contrast T1 weighted, T2/FLAIR) and quantitative MR imaging (Dynamic Contrast Enhanced (DCE) MRI, and Diffusion Weighted Imaging (DWI)). We use data from 39 patients from the Ivy Glioblastoma Atlas Project (Ivy GAP)[4] which provides matched imaging, ISH, RNA, gene expression and clinical data over the course of treatment. We analyze pre-operative anatomic imaging data to determine the tumor-induced mass effect in each patient using quantitative measures such as ‘Lateral ventricle displacement’ [2]. Perfusion and diffusion measures are derived from pre-operative DCE and DWI imaging. Additionally, we estimate IFF velocities in the tumor region using DCE imaging data in combination with a computational model of fluid flow [5].

Abstract IA09: Metabolic recycling in cancer

Abstract Metabolic reprogramming is a hallmark of cancer that supports the rapid proliferation and survival of tumor cells. While many studies have focused on identifying pathways involved in increased fuel usage, such as glucose and glutamine uptake and metabolism, the fate of metabolic waste products has been little investigated. Ammonia is a ubiquitous by-product of cellular metabolism, long thought to be a toxic waste product. Here we will discuss ammonia recycling to support cellular biomass and proliferation in breast cancer. Citation Format: Marcia Haigis. Metabolic recycling in cancer [abstract]. In: Proceedings of the AACR Special Conference: Advances in Modeling Cancer in Mice: Technology, Biology, and Beyond; 2017 Sep 24-27; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(10 Suppl):Abstract nr IA09.

The role of HIF-1α in chemo-/radioresistant tumors.

Chemo-/radioresistance is a major obstacle in clinical oncology. The precise failure mechanisms of chemo-/radioresistance are multifactorial failures. It is now widely accepted that a tumor hypoxia microenvironment contributes significantly to chemo-/radioresistance. Hypoxia is the most common and obvious neoplastic microenvironment and is due to the rapid proliferation of tumor cells. HIF-1α is a principal molecular mediator of adaptability to hypoxia in tumor cells. HIF-1α activation leads to the transcription of a plethora of target genes that promote physiological changes associated with chemo-/radioresistance, including increasing the ability of DNA repair, the inhibition of apoptosis, and alterations of the cellular metabolism. Moreover, recent findings suggest that HIF-1α-activated autophagy is a crucial factor in the promotion of cell survival under the distressed microenvironment, thereby leading to the chemo-/radioresistance. This chapter presents an overview of the role of HIF-1α in chemo-/radioresistance of tumor cells.

Beneficial Metabolic Consequences of Acyl‐CoA Synthetase ACSVL3 Knockout in Glioma Cells

We previously reported that knockdown or knockout (KO) of the fatty acid metabolic enzyme Very Long Chain Acyl‐CoA Synthetase 3 (ACSVL3; SLC27A3) in glioblastoma cells was beneficial. KO U87 glioma cells grew at a slower rate, became adherence‐dependent, and produced fewer, slower‐growing subcutaneous and intracranial tumors when implanted in NOD‐SCID mice. To understand the mechanisms underlying these changes, we investigated several possibilities. We initially demonstrated that lack of ACSVL3 did not slow the synthesis of membrane phospholipids necessary for the rapid proliferation of tumor cells in culture. We then investigated the possibility that ACSVL3 KO affected either apoptosis or autophagy. However, no differences in these processes between control and KO U87 cells were found. We next looked at cell cycle, and found that KO produced significant changes. In non‐synchronized cells, control U87 cells were mainly in G0/G1, while ACSVL3 KO cells were mainly in S‐phase. Both control and KO cells synchronized by serum starvation started out mainly in G0/G1, but by 12 hours KO cells shifted significantly toward S‐phase. Identification of the cyclins and cyclin‐dependent kinases that may be involved is in progress. Proteomic and metabolomic profiling indicated that in addition to effects on lipid metabolism, ACSVL3 KO produced changes in glucose and energy metabolism. Although glucose transporters GLUT1 and GLUT3 protein levels were reduced by KO, cellular uptake of labeled 2‐deoxyglucose was unaffected. However, oxidation of glucose to CO2 was reduced 40% by ACSVL3 depletion, and the cellular glucose level was 25% higher in KO cells. Glycolytic enzymes were upregulated by KO, but metabolic intermediates were essentially unchanged. Interestingly, lactate production and the levels of both LDHA and LDHB were elevated by ACSVL3 KO, and pentose phosphate pathway activity was lower in KO cells. Citric acid cycle enzymes, electron transport chain complexes, and ATP synthase protein levels were all reduced by ACSVL3 depletion. Mitochondria had a more punctate morphology in U87 cells, but were more elongated in KO cells. In conclusion, depletion of ACSVL3 affects many metabolic processes not directly related to lipid metabolism in human glioblastoma cells.Support or Funding InformationKennedy Krieger Research Administration fundsThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Abstract 1957: Molecular pathogenesis and drug synergism in a zebrafish model of high risk neuroblastoma

Abstract We have developed a transgenic zebrafish model that overexpresses MYCN and harbors loss-of-function mutations of the nf1 tumor suppressor. In this model, loss of nf1 leads to aberrant activation of RAS-MAPK signaling, promoting both increased tumor cell survival and rapid tumor cell proliferation. These neuroblastomas are very aggressive in that almost all of the fish develop neuroblastoma by 3 weeks of age. Three-week old juvenile fish are very small, making it feasible to test the effectiveness of many drugs and drug combinations in vivo for activity against the primary tumors. We demonstrate these advantages of the model by showing marked synergistic anti-tumor effects of a MEK inhibitor (trametinib) and a retinoid (isotretinoin) in vivo at several different dosage combinations by in vivo isobologram analysis. Thus, inhibition of RAS-MAPK signaling can significantly improve the treatment of this very aggressive form of neuroblastoma when it is combined with the inhibition of other key pathways. Because of the very high penetrance and rapid onset of neuroblastoma in our nf1-deficient, MYCN-transgenic zebrafish model, it is one of the only model systems in which extensive analysis of the synergistic activity of two or more drugs can be evaluated in primary tumors in vivo. This capability is especially valuable given that mutations causing RAS-MAPK pathway hyperactivation have been shown to arise frequently at the time of relapse of childhood neuroblastomas, indicating the need to eliminate these mutated tumor cells as a component of the primary treatment. Note: This abstract was not presented at the meeting. Citation Format: Shuning He, Marc R. Mansour, Mark W. Zimmerman, Hillary M. Layden, A. Thomas Look. Molecular pathogenesis and drug synergism in a zebrafish model of high risk neuroblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1957. doi:10.1158/1538-7445.AM2017-1957