Targeting Ferroptosis to Iron Out Cancer

Ferroptosis is a form of oxidative, iron-dependent, non-apoptotic cell death characterized by the accumulation of lipid peroxides and the depletion of glutathione. Ferroptosis plays a significant role in human cancer and is essential in neurological disorders, including neurodegeneration, stroke, and neurotrauma. One of the key challenges in cancer research is how to effectively kill cancer cells while leaving healthy cells intact. Cancer cells often have defects in cell death executioner mechanisms, which is one of the main reasons for therapy resistance. To enable growth, cancer cells exhibit an increased iron demand compared with normal, non-cancer cells. This iron dependency can make cancer cells more vulnerable to iron-catalyzed necrosis, referred to as ferroptosis. It is a newly identified regulated form of cell death, which is thought to play a major role in neurodegenerative diseases. The mechanisms of ferroptosis in several neurological disorders are discussed in detail in this article. It also provides an overview of emerging medications that target ferroptosis in the treatment of neurological disorders. It also highlights the variations and connections between the different cell death pathways implicated in neurological disorders. Clarifying the function of ferroptosis in the brain will help us better understand the mechanisms behind neurological disorders and offer possible strategies for both acute and long-term neurological illness prevention and therapy. Consequently, we provide an overview and brief description of the main pathways involved in ferroptosis in this review, focusing on its regulation and its dual roles as a tumor suppressor and an oncogenic process in various human malignancies. The identification of FDA-approved drugs as ferroptosis inducers has raised high expectations for ferroptosis as a promising new approach to killing therapy-resistant cancers.

Selective Cancer Targeting via Aberrant Behavior of Cancer Cell-associated Glucocorticoid Receptor

The challenges translating glioblastoma therapies lie in safeguarding normal neuronal and glial tissue. Considering that the central nervous system has the lowest regenerative capacity, the exclusive focus on tumor eradication while neglecting normal tissue response, impedes proper healing. This oversight leads to toxicities ultimately culminating in patient demise with marginal gain in survival. BPM31510 is a nanodispersion encapsulating highly hydrophobic oxidized Coenzyme Q10 (CoQ10), allowing for delivery of supraphysiological CoQ10 concentrations into cancer cells. Initial investigations demonstrated differential responsiveness between cancer and non-cancer cells in vitro, an effect associated with high superoxide levels exclusively in cancer cells. To evaluate this divergence between cancer and non-cancer cells, a competitive co-culture cell-based assay was developed with labeled pairs of glioblastoma and non-cancer cells incubated up to three weeks with various concentrations of BPM31510 to ascertain optimal concentration leading to preferential growth of non-cancer cells. Human astrocyte (HA), mouse fibroblast (NIH 3T3), and rat astrocyte (TNC1) cells were paired with U251 human glioma, mouse GL261 and rat C6 glioma, respectively. Cell growth and spatial extent of the plate containing non-cancer or tumor cells were monitored every other day. Using doxorubicin as control, no dosage was discerned in which non-cancer cells exhibited advantage over tumor cells (i.e., either cultures became all tumor cells or both cell lines were eradicated). Using various concentrations of BPM31510, we identified an optimal dose in which cancer cells diminished while non-cancer cells predominated, reflected by markedly elevated mitochondrial superoxide levels in cancer cells prior to cell death. A co-culture cell-based assay was developed to identify an optimal dose of BPM31510 with selective preservation of non-cancer cells alongside tumor cell death. This approach elucidates developing a “fine tuning” strategy, wherein BPM31510 is titrated over time, thereby promoting normal tissue and healing, and ultimately leading to superior and prolonged functional outcomes.

Selenium chemoprevention by apoptosis has been well studied, but it is not clear whether selenium can activate early barriers of tumorigenesis, namely senescence and DNA damage response. To test this hypothesis, we treated normal and cancerous cells with a gradient concentration of sodium selenite, methylseleninic acid and methylselenocysteine for 48 h, followed by a recovery of 1-7 days. Here we show that selenium compounds at doses of </=LD(50) can induce cellular senescence, as evidenced by the expression of senescence-associated beta-galactosidase and 5-bromo-2-deoxyuridine incorporation, in normal but not cancerous cells. In response to clastogens, the ataxia telangiectasia mutated (ATM) protein is rapidly activated, which in turn initiates a cascade of DNA damage response. We found that the ATM pathway is activated by the selenium compounds, and the kinase activity is required for the selenium-induced senescence response. Pretreatment of the MRC-5 non-cancerous cells with the antioxidant N-acetylcysteine or 2,2,6,6-tetramethylpiperidine-1-oxyl suppresses the selenium-induced ATM activation and senescence. Taken together, the results suggest a novel role of selenium in the activation of early tumorigenesis barriers specific in non-cancerous cells, whereby selenium induces an ATM-dependent senescence response that depends on reactive oxygen species.

Many quinoline-based compounds possess pharmacological properties that are beneficial for the control of various human diseases, including microbial infections, inflammatory-related disorders, cardiovascular conditions, and cancer. The anticancer activities of quinoline compounds are often rendered by their property of inhibiting protein kinases, proteasome activity, tubulin polymerization/depolymerization and DNA repair. To further improve the efficacy and specificity, we created and characterized several quinoline derivatives, which led us to identify CTRI-17 and CTRI-20 as promising leads. We examined their anti-proliferative properties using three human breast cancer cell lines (MDA-MB-468, MDA-MB-231 and MCF-7) and two matching non-cancer breast cell lines (184B5 and MCF10A). In addition, we also examined their efficacy using one leukemic cell line (K-562) and one cervical cell line (HeLa). Data from this study showed that the IC50 values of CTRI-17 and CTRI-20 were 0.1-0.3 μM and 1.2-2.4 μM ranges on cancer and non-cancer cell lines, respectively. Thus, both CTRI-17 and CTRI-20 killed cancer cells 10-20 times more effectively than non-cancer cells. This differential cell killing effects on cancer and non-cancer cells is a highly desirable property for potential anticancer therapeutics. We further found that cancer cells treated with the two compounds resulted in increases of pro-apoptotic proteins such as p53 and Bax, and decreases of anti-apoptotic proteins such as Bcl-2 and survivin. Our data obtained from flow cytometry and confocal microscopy showed that CTRI-17 and CTRI-20 induced prolonged cell cycle arrest at G2/M phase prior to causing apoptosis in cancer cells. However, the two compounds neither induced substantial cell cycle arrest nor caused high levels of apoptosis in non-cancer cells at the same drug concentration. We are currently in the process of determining the detailed functional mechanisms of CTRI-17 and CTRI-20. Citation Format: Indeewari K. Lindamulage, Hai-Yen Vu, Yi-Fang Lee, Hoyun Lee, Piyush Trivedi. Characterization of two novel quinoline derivatives that induce apoptosis in a cancer-specific manner. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 701. doi:10.1158/1538-7445.AM2013-701

Biosensor device for the photo-specific detection of immuno-captured bladder cancer cells using hexaminolevulinate: An ex-vivo study.

We previously presented data on the anticancer activity of a small library of synthetic flavonoid derivatives against glioblastoma and breast cancer lines in addition to several other human cancer cell lines. This data suggested that the chalcone structural backbone afforded more potent anticancer activity compared to its flavanone counterpart. Using this data, we selected three lead compounds: RK6, RK7, and RK15, to screen for activity against non‐cancerous cells to test for selectivity. Here we present updated IC50 values of these three analogs against MCF7 breast adenocarcinoma cells in addition to the previously reported data on the A‐172 glioblastoma cell line. To determine the selectivity of these lead compounds, RK6, RK7, and RK15 were screened against two non‐cancerous human cell lines: normal human astrocytes (NHA) and normal human mammary epithelial cells (NHME).The activity of our lead compounds against A‐172 and MCF7 cells were assessed using the XTT cell viability assay to determine respective IC50 values. As reported previously, the IC50 values of our lead compounds in the A‐172 cell line were 20 μM, 22 μM, and 3 μM for RK6, RK7, and RK15 respectively. Updated IC50 values of the compounds in the MCF7 cell line were 19 μM, 99 μM, and 12 μM for RK6, RK7, and RK15.In order to evaluate the selectivity of the lead compounds for cancerous vs. non‐cancerous cells, we utilized the Vialight cell viability assay to screen the chalcones in NHA and NHME cells. In the NHA cells, the determined IC50 values of RK6, RK7, and RK15 were 47 μM, 52 μM, and &gt;200 μM respectively. IC50 values were 60 μM, &gt;200 μM, and &gt;200 μM for the NHME cells treated with RK6, RK7, and RK15 respectively. This data was used to calculate selectivity indices for our lead compounds against A‐172 cells compared to NHA cells and MCF7 cells compared to NHME cells. For RK6, there was a 2.3 fold selectivity for A‐172 cells and a 3.1 fold selectivity for MCF7 cells compared to their non‐cancerous comparator cells. The selectivity index for RK7 showed a 2.3 fold selectivity for A‐172 compared to the NHA. In these instances, RK6 and RK7 appear to be somewhat selective for cancerous vs. non‐cancerous cell lines. Given the IC50 values &gt;200 μM for RK7 in NHA cells and RK15 in NHA and NHME cells, exact selectivity indices could not be determined. However, in these occurrences, it would appear that the compounds demonstrated a high degree of selectivity for cancerous vs. non‐cancerous cells.Collectively, these results suggest that our lead chalcone compounds are at least somewhat selective for cancerous vs. non‐cancerous cells. Interestingly, the most potent compound in the A‐172 and MCF7 cells, RK15, demonstrated the highest degree of selectivity. This is intriguing given that recent trends in anticancer drug discovery favor compounds that are selective for cancer cells. These findings may provide some insights into the yet to be elucidated mechanisms of cell death by these novel chalcone derivatives, and will form the basis of future studies.Support or Funding InformationThis work was supported in part by an AFPE Gateway Scholarship for the first author.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

The insulin-like growth factor (IGF) pathway represents one of the most studied molecular regulatory networks in oncology. Clinical trials investigating the therapeutic value of anti-IGF1 receptor (IGF1R) therapies in cancer, including prostate cancer, are ongoing. However, the multiple functions of the IGF network in the prostate are not entirely known. To elucidate the effects of IGF and insulin (INS) on prostate cells, we stimulated prostate cancer (PC3, DU145, LNCaP, DUCaP) and noncancerous prostate cells (EP156T, RWPE-1) and observed differing responses: whereas cancer cells responded to IGF and INS exposure by way of enhanced cell proliferation and glucose consumption, basal to luminal differentiation was induced in noncancerous cells. The same diverse responses were observed when the growth factor receptors IGF1R or INSR were overexpressed. Down-regulation of IGF1R or INSR isoform A (INSRA) also inhibited only proliferation of cancer cells. The proliferative response induced by the INSR in cancer cells was mediated solely by the INSRA. Moreover we observed that the receptors of the IGF network mutually influence their expression and exert redundant functions, thus underscoring the functional molecular network formed by IGF, INS, IGF1R, and INSR. Collectively we found that both IGF1R and INSRA have oncogenic effects in prostate cancer, but the IGF network also has important physiological functions in the noncancerous prostate. These data provide new insights into the biology of the IGF network in the prostate, thereby facilitating the design and interpretation of clinical studies investigating IGF1R targeting agents.

Background: The mechanisms underlying low intensity ultrasound (LIUS) mediated suppression of inflammation and tumorigenesis remain poorly determined.Methods: We used microarray datasets from NCBI GEO Dataset databases and conducted a comprehensive data mining analyses, where we studied the gene expression of 299 cell death regulators that regulate 13 different cell death types (cell death regulatome) in cells treated with LIUS.Results: We made the following findings: (1) LIUS exerts a profound effect on the expression of cell death regulatome in cancer cells and non-cancer cells. Of note, LIUS has the tendency to downregulate the gene expression of cell death regulators in non-cancer cells. Most of the cell death regulator genes downregulated by LIUS in non-cancer cells are responsible for mediating inflammatory signaling pathways; (2) LIUS activates different cell death transcription factors in cancer and non-cancer cells. Transcription factors TP-53 and SRF- were induced by LIUS exposure in cancer cells and non-cancer cells, respectively; (3) As two well-accepted mechanisms of LIUS, mild hyperthermia and oscillatory shear stress induce changes in the expression of cell death regulators, therefore, may be responsible for inducing LIUS mediated changes in gene expression patterns of cell death regulators in cells; (4) LIUS exposure may change the redox status of the cells. LIUS may induce more of antioxidant effects in non-cancer cells compared to cancer cells; and (5) The genes modulated by LIUS in cancer cells have distinct chromatin long range interaction (CLRI) patterns to that of non-cancer cells.Conclusions: Our analysis suggests novel molecular mechanisms that may be utilized by LIUS to induce tumor suppression and inflammation inhibition. Our findings may lead to development of new treatment protocols for cancers and chronic inflammation.

Diets enriched with bioactive food components trigger molecular changes in cells that may contribute to either health-promoting or adverse effects. Recent technological advances in high-throughput data generation allow for observing systems-wide molecular responses to cellular perturbations with nontoxic and dietary-relevant doses while considering the intrinsic differences between cancerous and noncancerous cells. In this chemical profile, we compared molecular responses of the colon cancer cell line HT29 and a noncancerous colon epithelial cell line (HCEC) to two widely encountered food components, sulforaphane and selenium. We conducted this comparison by generating new transcriptome data by microarray gene-expression profiling, analyzing them statistically on the single gene, network, and functional pathway levels, and integrating them with protein expression data. Sulforaphane and selenium, at doses that did not inhibit the growth of the tested cells, induced or repressed the transcription of a limited number of genes in a manner distinctly dependent on the chemical and the cell type. The genes that most strongly responded in cancer cells were observed after treatment with sulforaphane and were members of the aldo-keto reductase (AKR) superfamily. These genes were in high agreement in terms of fold change with their corresponding proteins (correlation coefficient r(2) = 0.98, p = 0.01). Conversely, selenium had little influence on the cancer cells. In contrast, in noncancerous cells, selenium induced numerous genes involved in apoptotic, angiogenic, or tumor proliferation pathways, whereas the influence of sulforaphane was very limited. These findings contribute to defining the significance of cell type in interpreting human cellular transcriptome-level responses to exposures to natural components of the diet.

Differential Allele-Specific Expression Uncovers Breast Cancer Genes Dysregulated by Cis Noncoding Mutations.

Despite recent notable progress in medical sciences, cancer remains a leading cause of human death worldwide, accounting for about one-quarter of human deaths in wealthy countries and about one-eighth worldwide (Ferlay et al. 2010). In fact, the progress in cancer research has been slower compared with that achieved related to other pathologies, such as cardiovascular disorders. This is mainly because of the enormous complexity of this disease, which exhibits sophisticated cellular mechanisms that are the targets of evolutionary processes driven by random genetic and epigenetic mutations.

A Competitive Co-cultivation Assay for Cancer Drug Specificity Evaluation

Dietary factors play an important role in both the development and prevention of human cancers, including breast carcinoma. One dietary micronutrient, sodium butyrate (NaB), is a major end product of dietary starch and fiber, produced naturally during digestion by anaerobic bacteria in the cecum and colon. NaB is a potent growth inhibitor and initiates cell differentiation for many cell types in vitro. In this study, we investigated the effects of NaB on three human mammary epithelial cells and regulation of the IGF axis, specifically, IGF-binding protein-3 (IGFBP-3), a known growth regulator in human mammary cells, and IGFBP-related protein 2 (IGFBP-rP2)/connective tissue growth factor. NaB inhibited DNA synthesis, as measured by [3H]thymidine incorporation, in estrogen-responsive (MCF-7) and estrogen-non-responsive (Hs578T) breast cancer cells, and normal human mammary epithelial cells (HMEC) to a similar degree (up to 90% inhibition at 1-10 mM concentrations). Treatment of cells with NaB induced histone hyperacetylation, suggesting that NaB exerts its biological effects, at least in part, as a histone deacetylase inhibitor in mammary epithelial cells. Treatment of Hs578T cells with NaB caused an induction of apoptotic cell death. NaB treatment resulted in increased levels of p21(Waf1/Cip1) mRNA and protein in Hs578T cells and distinct upregulation of p27(Kip1) in HMEC, suggesting that NaB activates different genes involved in cell cycle arrest, depending upon the cell type. In the same context, among the IGFBP superfamily members tested, NaB specifically upregulated the expression of IGFBP-3 and IGFBP-rP2. These two proteins are known to be involved in inhibition of mammary epithelial cell replication. Northern blot analysis showed that NaB treatment at 1-10 mM concentrations caused a dose-dependent stimulation of IGFBP-3 mRNA expression in cancerous cells and IGFBP-rP2 mRNA expression in both cancerous and non-cancerous cells. Protein data from Western ligand blot and immunoblot analyses demonstrated parallel results. In summary, we have demonstrated that NaB (i) uniformly suppresses DNA synthesis in both cancerous and non-cancerous mammary cells, and (ii) upregulates IGFBP-3 and IGFBP-rP2 mRNA and protein levels in cancerous and non-cancerous mammary cells. These results provide the first demonstration that butyrate regulates the IGFBP system in the human mammary system.

Cancer is the leading cause of death worldwide and all age groups, including children, are at risk for cancer associated death. Disease progression and metastasis are major contributors to cancer-associated morbidity and mortality. Increased cell migration rate is characteristic of tumor progression and metastasis. Actin binding proteins regulate cytoskeletal remodeling at the leading edges of cancer cells, facilitate invasive organelle (invadopodia) formation, and promote higher migration rates. Docosahexaenoic acid (DHA), a poly unsaturated fatty acid, has been shown to inhibit the cancer cell metastatic phenotype. Here, we test the hypothesis that changes in actin binding proteins regulate cancer cell migration and that supplementing cells with DHA will prevent these changes. Non-cancer (MLE12) and cancer (A549, HCT116 and MCF7) cells were treated with 8-Br-cAMP and/or DHA. F-actin content was measured using confocal microscopy. Cell migration was estimated by wound assay and transwell apparatus. Actin binding proteins, profilin, cofilin, vimentin and gelsolin, were identified and quantified using confocal microscopy and western blot to evaluate wound edges and actin co- munoprecipitate, respectively. F-actin content and cell migration were increased by cAMP in association with changes in profilin, cofilin, vimentin and gelsolin levels. DHA treatment suppressed the increase in actin content and cell migration in cancer cells but not in non-cancer cells in association with altered levels of actin binding proteins. We postulate that DHA specifically inhibits cancer cell migration via alterations in actin binding proteins indicating a therapeutic potential against cancer cell metastasis. The changes in actin binding proteins could serve as biomarkers for cancer progression and as innovative therapeutic targets. Citation Format: Mehboob Ali, Alexander Long, Kathryn M. Heyob, Asha Srinivasan, Lynette K. Rogers. Modulation of actin binding proteins by docosahexaenoic acid reduces cancer cell migration. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4057. doi:10.1158/1538-7445.AM2014-4057