Reduced Cancer Cell Viability Research Articles

Knockdown of DJ-1 Resulted in a Coordinated Activation of the Innate Immune Antiviral Response in HEK293 Cell Line

PARK7, also known as DJ-1, plays a critical role in protecting cells by functioning as a sensitive oxidation sensor and modulator of antioxidants. DJ-1 acts to maintain mitochondrial function and regulate transcription in response to different stressors. In this study, we showed that cell lines vary based on their antioxidation potential under basal conditions. The transcriptome of HEK293 cells was tested following knockdown (KD) of DJ-1 using siRNAs, which reduced the DJ-1 transcripts to only 12% of the original level. We compared the expression levels of 14k protein-coding transcripts and 4.2k non-coding RNAs relative to cells treated with non-specific siRNAs. Among the coding genes, approximately 200 upregulated differentially expressed genes (DEGs) signified a coordinated antiviral innate immune response. Most genes were associated with the regulation of type 1 interferons (IFN) and the induction of inflammatory cytokines. About a quarter of these genes were also induced in cells treated with non-specific siRNAs that were used as a negative control. Beyond the antiviral-like response, 114 genes were specific to the KD of DJ-1 with enrichment in RNA metabolism and mitochondrial functions. A smaller set of downregulated genes (58 genes) was associated with dysregulation in membrane structure, cell viability, and mitophagy. We propose that the KD DJ-1 perturbation diminishes the protective potency against oxidative stress. Thus, it renders the cells labile and responsive to the dsRNA signal by activating a large number of genes, many of which drive apoptosis, cell death, and inflammatory signatures. The KD of DJ-1 highlights its potency in regulating genes associated with antiviral responses, RNA metabolism, and mitochondrial functions, apparently through alteration in STAT activity and downstream signaling. Given that DJ-1 also acts as an oncogene in metastatic cancers, targeting DJ-1 could be a promising therapeutic strategy where manipulation of the DJ-1 level may reduce cancer cell viability and enhance the efficacy of cancer treatments.

Effects of Lactococcus lactis on colorectal cancer in various terms: a narrative review

Introduction: Colorectal cancer is one of the most common cancers with an increasing number of cases. Various studies have found an association between the gut microbiota balance and colorectal cancer incidence. Lactococcus lactis is a probiotic bacterium found in fermented foods, particularly yogurt and cheese. This probiotic has been shown to reduce various anti-inflammatory and pro-inflammatory agents that trigger cancer, such as interleukin (IL)-6, IL-18, tumour necrosis factor-alpha (TNF-α), and natural killer (NK) cells. Methods: Full-text articles and original research published in the last ten years were used as references, and “Lactococcus and colorectal cancer” as keywords. The reference search is on several databases, such as PubMed, ScienceDirect, ProQuest, and Nature. Searching results obtained eleven articles. Discussion: Lactococcus lactis does have a perfect role in suppressing cancer cells. Lactococcus lactis has anti-proliferative effects associated with decreased cyclin D1 expression in SW480 cell lines, decreased NK cells, reduced cancer cell viability, decreased IL-8 levels, and decreased IL-6. Conclusion: Lactococcus lactis contains nisin, which can suppress various gene, protein, and cytokine expressions that play a role in cancer cell growth. Probiotics can inhibit colorectal cancer without significant side effects.

PI3K inhibitor “alpelisib” alleviates methotrexate induced liver injury in mice and potentiates its cytotoxic effect against MDA-MB-231 triple negative breast cancer cell line

Hepatotoxicity is the main off-target effect of methotrexate (MTX) limiting its effective clinical use. Besides, MDA-MB231 breast cancer cells show chemoresistance, partly via PI3K/AKT pathway. Therefore, we investigated the ameliorative potentials of the PI3K inhibitor, alpelisib (ALP) on MTX-induced hepatotoxicity (in vivo) and the restraining potentials of ALP on MDA-MB231 chemoresistance to MTX (in vitro). Twenty-eight male BALB/c mice were divided into 4 groups. In treatment groups, mice were administered ALP (2.5 and 5 mg/kg) for 5 days and MTX (20 mg/kg) from day 2 till day 5. The results showed that ALP restored hepatic architecture, reduced immune cell infiltration (F4/80, Ly6G and MPO) and repressed the rise in liver enzymes (AST and ALT) induced by MTX. Additionally, ALP rectified the MTX-induced disruption of cellular oxidant status by boosting antioxidant defense systems (HO-1 and GSH) and repressing lipid peroxidation (MDA and 4-HNE). Finally, ALP curbed MTX-induced hepatocyte apoptosis (NF-κB and BAX) and shifted the cytokine milieu away from inflammation (IL-17, IL-22, IL-6 and IL- 10). The results of the in vitro experiments revealed that ALP alone and in combination with MTX, synergistically, reduced cancer cell viability (MTT assay), migration (wound healing assay) and their capacity to establish colonies (colony formation assay) as compared to MTX alone. RT-PCR revealed the antiproliferative (Bcl-2) and proapoptotic (BAX) potentials of ALP and ALP/MTX combination especially after 24 h. In conclusion, targeting PI3K/AKT pathway is a promising strategy in triple negative breast cancer patients by ameliorating hepatotoxicity and restraining chemoresistance to chemotherapy.

Structural investigation of interactions between halogenated flavonoids and the lipid membrane along with their role as cytotoxic agents

This study focuses on understanding the structural and molecular changes in lipid membranes under the influence of six halogenated flavonoid derivatives differing in the number and position of substitution of chlorine and bromine atoms (D1–D6). Utilizing various analytical techniques, including fluorometric methods, dynamic light scattering (DLS), attenuated Fourier transform infrared spectroscopy (ATR- FTIR), and FT-Raman spectroscopy, the research aims to elucidate the mechanisms underlying the interaction of flavonoids with cell membranes. Additionally, the study includes in silico analyses to explore the physicochemical properties of these compounds and their potential pharmaceutical applications, along with toxicity studies to assess their effects on cancer, normal, and red blood cells. Our study showed the ability of halogenated derivatives to interact mostly with the outer part of the membrane, especially in the lipid heads region however, some of them were able to penetrate deeper into the membrane and affect the fluidity of hydrocarbon chains. The potential to reduce cancer cell viability, the lack of toxicity towards erythrocytes, and the favourable physicochemical and pharmacokinetic properties suggest these halogenated flavonoids potential candidates for exploring their potential for medical use.

Navigating the landscape of epitranscriptomics and host immunity.

RNA modifications, also termed epitranscriptomic marks, encompass chemical alterations to individual nucleotides, including processes such as methylation and editing. These marks contribute to a wide range of biological processes, many of which are related to host immune system defense. The functions of immune-related RNA modifications can be categorized into three main groups: regulation of immunogenic RNAs, control of genes involved in innate immune response, and facilitation of adaptive immunity. Here, we provide an overview of recent research findings that elucidate the contributions of RNA modifications to each of these processes. We also discuss relevant methods for genome-wide identification of RNA modifications and their immunogenic substrates. Finally, we highlight recent advances in cancer immunotherapies that aim to reduce cancer cell viability by targeting the enzymes responsible for RNA modifications. Our presentation of these dynamic research avenues sets the stage for future investigations in this field.

Anti-cancer Effect of Neocuproine on Oxidative Stress and Inflammatory Levels Induced by Neuroblastoma Cancer Cells

Neocuproine is a free radical scavenger. Neuroblastoma cancer is seen mostly in children. It accounts for approximately 15% of all pediatric cancer deaths. Treatments against SH-SY5Y may be ineffective due to the development of drug resistance, resulting in poor survival in high-risk patients. The goal of this study was to find out how the anti-cancer concentration of Neocuproine affected the levels of oxidants, antioxidants, and cytokines in the SH-SY5Y cell line. Five different concentrations of Neocuproine (12.5, 25, 50, 100, and 200 µM) were applied to the cell line for 24 h. MTT cytotoxicity tests, total oxidant status and total antioxidant capacity tests, and anti-inflammatory cytokine tests were used to look at the uses of Neocuproine. In our results, 100 µM Neocuproine application reduced cancer cell viability up to 69.76%. In addition, it was determined that 100 µM Neocuproine reduced oxidant activity, increased antioxidant capacity, and regulated anti-inflammatory cytokine levels in neuroblastoma cells, consistent with the cytotoxicity test. As a result, it was concluded that Neocuproine has an anti-cancer effect on the SH-SY5Y cell line.

Abstract LB296: Human CYP1B1 enhances cancer progression through induction of Sp1 via DNMT-mediated epigenetic regulation of miR-375

Abstract Human cytochrome P450 1B1 (CYP1B1) is a major catalytic enzyme for metabolism of 17β-estradiol (E2) to catechol estrogens, mainly carcinogenic metabolite 4-hydroxyestradiol (4-OHE2). To identify the unknown molecular mechanism for cancer progression induced by CYP1B1, we studied the role of CYP1B1 in human tumor cells, MCF-7, MDA-MB-231, and HeLa cells. Suppression of CYP1B1 by shRNA significantly reduced cancer cell viability and invasion. Clinical data from cancer patients showed that high level of CYP1B1 expression resulted in relatively lower survival rate. Interestingly, overexpression of CYP1B1 markedly activated Wnt/β-catenin signaling and EMT process, and suppression of specificity protein 1 (Sp1) by siRNA, an well-known transcription factor, demolished the promoting effects of CYP1B1 on cancer progression. To investigate the correlation between CYP1B1 and Sp1, we analyzed the clinical data from breast and cervical cancer patients and the results showed the positive correlation between two genes. Furthermore, TMS (tetramethoxystilbene), a specific CYP1B1 inhibitor, diminished the induction of Sp1 by CYP1B1, and 4-OHE2 exceedingly promoted the expression level of Sp1. Surprisingly, miR-375, known as an oncogenic microRNA, substantially decreased by 4-OHE2 and CYP1B1 overexpression in CYP1B1-transgenic (TG) mice. Methylation specific assay revealed that CYP1B1 suppressed miR-375 expression through induction of hyper-methylation on pre-miR-375. In addition, epigenetic regulation of miR-375 by CYP1B1 was executed via induction of DNA methyltransferases (DNMTs), DNMT1, 3a, and 3b. Enhanced expression level of Sp1 by 4-OHE2 was significantly deteriorated by 5-aza-dC, a DNMT inhibitor. Taken together, our data suggest that CYP1B1 enhances cancer progression through induction of cancer cell proliferation and invasion via DNMT-mediated epigenetic regulation of miR-375 and further promotion of Sp1 expression. Citation Format: Young-Jin Chun, Yeo-Jung Kwon, Tae-Uk Kwon. Human CYP1B1 enhances cancer progression through induction of Sp1 via DNMT-mediated epigenetic regulation of miR-375 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB296.

Triple Negative Breast Cancer (TNBC): Cannabis sativa-Role of Phytocannabinoids

This review paper highlights the role of phytocannabinoids in controlling triple-negative breast cancer (TNBC). Triple-negative breast cancer (TNBC) is a specific subtype of breast cancer that does not express estrogen receptor (ER), progesterone receptor (PR), or human epidermal growth factor receptor 2 (HER-2), which has clinical features that include high invasiveness, high metastatic potential, proneness to relapse, and poor prognosis. TNBC is one of the most aggressive subtypes of breast cancer that is often associated with poor patient outcomes because of the development of metastases in secondary organisms like in the brain, bone, and lungs. Triple-negative breast cancer (TNBC) is a kind of breast cancer that does not have any of the receptors that are commonly found in breast cancer. The highest number of TNBC cases has been recorded in India. Currently, TNBC is highly prevalent among Indian women and develops approximately 20% to 43% of all patients with breast cancer. Triple-negative breast cancer (TNBC) is an aggressive malignancy that requires effective targeted drug therapy. Patients with TNBC develop metastasis and recurrence over time and have reduced survival compared to patients with other subtypes of breast cancer. Pre-clinical studies have demonstrated that phytocannabinoids exert important antitumor properties in the main breast cancer subtypes, particularly in TNBC, where different phytocannabinoids and synthetic cannabinoids have shown interesting therapeutic actions. Neither CBD nor THC are universally efficacious in reducing cancer cell viability. Focusing on in vitro studies, the effect of CBD on cancer cell viability ranges from no effect, to a modest reduction, and to significant cytotoxicity depending on concentrations, cancer cell lines, cell growth conditions, the performed assays, and the time of CBD exposure. However, it should also be noted that research into the efficacy, dosage and drug safety of cannabinoids in tumor therapy still has a long way to go, especially with regard to human clinical trials to be conducted, through which alone the benefits and advantages for cancer patients but also possible risks can be defined.

Neutron Capture Enhances Dose and Reduces Cancer Cell Viability in and out of Beam During Helium and Carbon Ion Therapy

Neutron capture enhanced particle therapy (NCEPT) is a proposed augmentation of charged particle therapy that exploits thermal neutrons generated internally, within the treatment volume via nuclear fragmentation, to deliver a biochemically targeted radiation dose to cancer cells. This work is the first experimental demonstration of NCEPT, performed using both carbon and helium ion beams with 2 different targeted neutron capture agents (NCAs). Human glioblastoma cells (T98G) were irradiated by carbon and helium ion beams in the presence of NCAs [10B]-BPA and [157Gd]-DOTA-TPP. Cells were positioned within a polymethyl methacrylate phantom either laterally adjacent to or within a 100 × 100 × 60 mm spread out Bragg peak (SOBP). The effect of NCAs and location relative to the SOBP on the cells was measured by cell growth and survival assays in 6 independent experiments. Neutron fluence within the phantom was characterized by quantifying the neutron activation of gold foil. Cells placed inside the treatment volume reached 10% survival by 2 Gy of carbon or 2 to 3 Gy of helium in the presence of NCAs compared with 5 Gy of carbon and 7 Gy of helium with no NCA. Cells placed adjacent to the treatment volume showed a dose-dependent decrease in cell growth when treated with NCAs, reaching 10% survival by 6 Gy of carbon or helium (to the treatment volume), compared with no detectable effect on cells without NCA. The mean thermal neutron fluence at the center of the SOBP was approximately 2.2 × 109 n/cm2/Gy (relative biological effectiveness) for the carbon beam and 5.8 × 109 n/cm2/Gy (relative biological effectiveness) for the helium beam and gradually decreased in all directions. The addition of NCAs to cancer cells during carbon and helium beam irradiation has a measurable effect on cell survival and growth in vitro. Through the capture of internally generated neutrons, NCEPT introduces the concept of a biochemically targeted radiation dose to charged particle therapy. NCEPT enables the established pharmaceuticals and concepts of neutron capture therapy to be applied to a wider range of deeply situated and diffuse tumors, by targeting this dose to microinfiltrates and cells outside of defined treatment regions. These results also demonstrate the potential for NCEPT to provide an increased dose to tumor tissue within the treatment volume, with a reduction in radiation doses to off-target tissue.

Exploring Anti-Breast Cancer Effects of Live Pediococcus acidilactici and Its Cell-Free Supernatant Isolated from Human Breast Milk.

Current breast cancer treatment options are limited by drug resistance and adverse side effects, which calls for the need for alternatives or complementary remedies. Probiotic bacteria isolated from human breast milk have been shown to possess proapoptotic and anti-inflammatory properties against breast mastitis in breastfeeding mothers and are being studied as possible anticancer regimens. Thus, this study aimed at exploring the effect of lactic acid bacteria isolated from human breast milk on MDA-MB 231 breast cancer cells. A total of twenty-two bacteria were isolated from four human breast milk samples. The isolates were characterized and identified using biochemical tests and Sanger sequencing, respectively. For in vitro experiments, we used isolated P. acidilactici to treat MDA-MB-231 cells, and an MTT assay was used to detect proliferation. RT-qPCR and wound healing assays were performed to determine the effect of the isolated P. acidilactici on breast cancer cytokine expression and migration. Exposure of MDA-MB 231 breast cancer cells to live P. acidilactici and its cell-free supernatant (CFS) for 24 h resulted in a reduction in cancer cell viability. Also, the expression of the cytokines IL-6, IL-8, and IL-10 in the breast cancer cells increased following exposure to P. acidilactici and its CFS for 24 and 72 h. Additionally, the levels of the SLUG gene remained unchanged while the TWIST1 gene was upregulated following exposure of the cancer cells to bacteria, indicating that P. acidilactici may promote epithelial-mesenchymal transition in breast cancer. Finally, the CFS significantly inhibited cancer cell mobility. These findings serve as a foundation to further investigate the usefulness of P. acidilactici as a potential therapeutic agent in breast cancer therapy.

Assessing the Therapeutic Impacts of HAMLET and FOLFOX on BRAF-Mutated Colorectal Cancer: A Study of Cancer Cell Survival and Mitochondrial Dynamics In Vitro and Ex Vivo.

Background and Objectives: Colorectal cancer (CRC) is a major global health challenge. The BRAF V600E mutation, found in 8-12% of CRC patients, exacerbates this by conferring poor prognosis and resistance to therapy. Our study focuses on the efficacy of the HAMLET complex, a molecular substance derived from human breast milk, on CRC cell lines and ex vivo biopsies harboring this mutation, given its previously observed selective toxicity to cancer cells. Materials and Methods: we explored the effects of combining HAMLET with the FOLFOX chemotherapy regimen on CRC cell lines and ex vivo models. Key assessments included cell viability, apoptosis/necrosis induction, and mitochondrial function, aiming to understand the mutation-specific resistance or other cellular response mechanisms. Results: HAMLET and FOLFOX alone decreased viability in CRC explants, irrespective of the BRAF mutation status. Notably, their combination yielded a marked decrease in viability, particularly in the BRAF wild-type samples, suggesting a synergistic effect. While HAMLET showed a modest inhibitory effect on mitochondrial respiration across both mutant and wild-type samples, the response varied depending on the mutation status. Significant differences emerged in the responses of the HT-29 and WiDr cell lines to HAMLET, with WiDr cells showing greater resistance, pointing to factors beyond genetic mutations influencing drug responses. A slight synergy between HAMLET and FOLFOX was observed in WiDr cells, independent of the BRAF mutation. The bioenergetic analysis highlighted differences in mitochondrial respiration between HT-29 and WiDr cells, suggesting that bioenergetic profiles could be key in determining cellular responses to HAMLET. Conclusions: We highlight the potential of HAMLET and FOLFOX as a combined therapeutic approach in BRAF wild-type CRC, significantly reducing cancer cell viability. The varied responses in CRC cell lines, especially regarding bioenergetic and mitochondrial factors, emphasize the need for a comprehensive approach considering both genetic and metabolic aspects in CRC treatment strategies.

Aptamer-modified tetrahedral DNA nanostructure-immobilized liposome for specific gene delivery and potential cancer theragnostic.

Targeted delivery of therapeutic agents to cancer cells is crucial for effective cancer treatment without adverse effects. In this study, we developed a novel delivery carrier, Aptamer-modified tetrahedral DNA nanostructure (TDN) immobilized Liposome (ApTL), for specific delivery to nucleolin-overexpressing cancer cells. We demonstrated that targeted ApTL was highly effective in delivering plasmid and mRNA to nucleolin-overexpressing cancer cells compared to non-targeted ApTL with a non-specific aptamer. ApTL, which is highly negative and nano-sized, specifically delivered nucleic acids to MDA-MB-231 and HeLa cancer cells, primarily via lipid-raft-mediated endocytosis. Furthermore, the co-delivery of mRNA and doxorubicin resulted in increased apoptosis and reduced cancer cell viability. Interestingly, co-delivery of mRNA and Dox did not show a significant difference in EGFP expression at 24h but dramatically increased EGFP expression at 48h, making ApTL/mEGFP/Dox a promising candidate for detecting live cancer cells after targeted cancer drug treatment. Our results suggest that ApTL can be a promising tool for the targeted delivery of therapeutic agents to nucleolin-overexpressing cancer cells, providing a new strategy for cancer theragnostic.

Plant based biosynthesis of TiO2NPs and evaluation of their cytotoxicity, antihemolytic, and protein antidenaturation effects

The current research reports the physicochemical and biological properties of biosynthesized titanium dioxide nanoparticles (TiO2NPs) using bioactive compounds of Achillea wilhelmsii C. Koch extract (Aw-TiO2NPs). The cytotoxic impacts of Aw-TiO2NPs were assessed by the MTT method on A375 and MCF-7 cancer cell lines and normal skin fibroblast cell line and also by hemolysis test against horse red blood cells (RBCs). In addition, apoptotic activity was analyzed by assaying the generation of reactive oxygen species (ROS) and reduction of mitochondrial membrane potential (MMP). Additionally, the potential of Aw-TiO2NPs in inhibiting protein denaturation was evaluated. The prepared Aw-TiO2NPs with a size range of 3–20 nm and almost spherical shape showed an anatase crystallographic phase. A significant dose-dependent reduction in cancer cell viability was observed against anatase Aw-TiO2NPs with IC50 values of 12.5 and 25 µg/mL for A375 and MCF-7 cells, respectively. Aw-TiO2NPs induced the generation of excessive ROS (95.9 %-215.4 %) and decreased MMP (60.9 %-72 %) in cancer cells, which leads to the initiation of intrinsic apoptosis. Furthermore, anatase Aw-TiO2NPs had no hemolytic effects on horse-RBCs and excellently inhibited protein denaturation (80.8 % at 500 µg/mL). Therefore, anatase Aw-TiO2NPs can be a valuable and biocompatible candidate for biomedical applications and clinical uses.

Green synthesis, characterization of melatonin-like drug bioconjugated CoS quantum dots and its antiproliferative effect on different cancer cells.

Quantum dots are usually particles smaller than 100nm and have a low toxic effect. This study aimed to bioconjugate the anticancer effective melatonin agonist to quantum dots and demonstrate its effects in two cancer lines. This is the first study that aims to examine the anticancer activity of ramelteon bioconjugation to quantum dots, providing a new perspective on the use of Melatonin and its derivatives in cancer. For this purpose, first of all, cobalt sulfide (CoS) quantum dots were synthesized, bioconjugated and characterized with Punica granatum extract by green synthesis method. The effects of synthesized nanomaterials on neuroblastoma and prostate cancer cells were investigated. It was noted that nanomaterials reduced cell viability by 50% in neuroblastoma and prostate cancer lines at a dose of 50µg/mL. Ramelteon bioconjugated nanomaterials reduced cancer cell viability by 1.4 times more than free melatonin. CoS quantum dots were determined to double the oxidative stress in the neuroblastoma cell line compared to the control, while no change was observed in prostate cancer. In the gene expression findings, it was observed that CoS nanoparticles caused an increase in the expression levels of apoptosis-related genes in the neuroblastoma cell line and induced key protein expression levels of pathways such as ROR-alpha in the prostate cancer cell line. As a result, it was observed that the viability of the neuroblastoma cell line decreased with apoptosis induced by oxidative stress, while this effect was observed in the DU-145 cell line via the ROR-alpha pathway.

New insights into the cytotoxic effects of Thymus vulgaris essential oil on the human triple-negative breast cancer cell line MDA-MB-231

Essential oils (EOs) are natural products that have gained wide interest due to their biological activities and anticancer properties through various mechanisms. The present study aimed to test the cytotoxicity of Thymus vulgaris L. (thyme) EO of Italian origin, rich in thymol (49.6%) and p-cymene (18.8%), towards the triple-negative breast cancer cell line MDA-MB-231 and to investigate the biochemical mechanisms underlying its antitumor activity. Thyme EO reduced cancer cell viability in a dose-dependent manner after 24 h treatment, with an IC50 value equal to 75.1 ± 15.2 μg/ml; simultaneously, the inhibition of cancer cell migration and colony formation capacity was evidenced. Thyme EO antiproliferative effects were related to the induction of apoptosis as demonstrated by the increased expression of the pro-apoptotic proteins Bax, cleaved caspase-3, phospho-p53, and SMAC/Diablo and by the reduction of the anti-apoptotic proteins Bcl-2, cIAP-1, cIAP-2, HIF-1α, survivin, and XIAP. Thyme EO administration led to the early formation of intracellular ROS, followed by the increment of MDA as an index of lipid peroxidation and by the decreased expression of the antioxidant enzymes catalase and PON2. The upregulation of Nrf2 mRNA expression and the strong induction of HO-1 sustained the activation of the Nrf2 pathway by thyme EO. These data showed that the EO from Thymus vulgaris L. might inhibit the malignant phenotype of MDA-MB-231, thus suggesting potential benefits against human triple-negative breast cancer.

NDUFA4 promotes the progression of head and neck paraganglioma by inhibiting ferroptosis.

NDUFA4 is a component of respiratory chain-oxidative phosphorylation pathway. NDUFA4 is highly expressed in tumor tissues, but little is known about the function of NDUFA4 in head and neck paraganglioma (HNPGL). We examined NDUFA4 expression in tissues from 10 HNPGL patients and 6 controls using qRT-PCR and Western blotting. NDUFA4 knockdown PGL-626 cells were established by using lentivirus infection and puromycin screening. Cell viability, ATP production, lipid reactive oxygen species, and mitochondrial membrane potential assays were performed to investigate the ferroptotic effects in NDUFA4 deficiency HNPGL cancer cells. Xenograft mouse model was created to detect the synergetic antitumor action between NDUFA4 deficiency and Metformin. NDUFA4 was upregulated in tumor tissues of HNPGL patients. NDUFA4 knockdown impaired the assembly of mitochondrial respiratory chain complexes and decreased the production of ATP and reduced cancer cell viability. Mechanistically, NDUFA4 knockdown increased cell ferroptosis, which further promoted Metformin-induced ferroptosis in PGL-626 cells. Therefore, NDUFA4 deficiency enhanced Metformin-mediated inhibition of the HNPGL progression in mice. In conclusion, NDUFA4 promotes the progression of HNPGL, and NDUFA4 knockdown enhances Metformin-mediated inhibition of the HNPGL progression in a mouse model.

Cytotoxicity of subcritical water extracts obtained from byproducts generated at commercial pecan shelling operations on cancer cells

This study examined potential of the extracts obtained from the byproducts generated at commercial pecan nut-shelling operations in cancer treatment. The subcritical water extracts obtained from two varieties, Native and Pawnee, were analyzed for their phenolic contents and compositions. Effects of the extracts on viability and IC50 of the human cell lines representing a broad range of cancer types, cervical, lung, skin, breast, colon and prostate cancers, were investigated. Although the effect of the temperature on the phenolic contents and compositions of the extracts was not statistically significant, the influence of the variety was extensive. The pecan shell extracts were not cytotoxic to the healthy cell line Vero in the concentration range examined. Some of the pecan shell extracts had greater efficay than Doxorubicin, a drug used in cancer chemotherapy, in reducing cancer cell viability. This study is novel and practical implications of the data generated in this study are noteworthy, because this is the first report on the beneficial effects of subcritical water extracts obtained from pecan shelling industry byproducts on a broad range of cancer cell lines. It is likely that the experimental data presented in this study will support and encourage future research on the biological pathways involved in the interactions of the cancer cells and the extracts. The findings of this study will facilitate research on downstream processing and purification of the crude extracts exhibiting high cancer cell cytotoxcity, potentially improving the final product efficacy and lead to commercial applications.Graphical

CLK2 and CLK4 are regulators of DNA damage-induced NF-κB targeted by novel small molecule inhibitors

Transcription factor NF-κB potently activates anti-apoptotic genes, and its inactivation significantly reduces tumor cell survival following genotoxic stresses. We identified two structurally distinct lead compounds that selectively inhibit NF-κB activation by DNA double-strand breaks, but not by other stimuli, such as TNFα. Our compounds do not directly inhibit previously identified regulators of this pathway, most critically including IκB kinase (IKK), but inhibit signal transmission in-between ATM, PARP1, and IKKγ. Deconvolution strategies, including derivatization and invitro testing in multi-kinase panels, yielded shared targets, cdc-like kinase (CLK) 2 and 4, as essential regulators of DNA damage-induced IKK and NF-κB activity. Both leads sensitize to DNA damaging agents by increasing p53-induced apoptosis, thereby reducing cancer cell viability. We propose that our lead compounds and derivatives can be used in context of genotoxic therapy-induced or ongoing DNA damage to increase tumor cell apoptosis, which may be beneficial in cancer treatment.

Dual CSF1R inhibition and CD40 activation demonstrates anti-tumor activity in a 3D macrophage- HER2+ breast cancer spheroid model.

The complex interaction between tumor-associated macrophages (TAMs) and tumor cells through soluble factors provides essential cues for breast cancer progression. TAMs-targeted therapies have shown promising clinical therapeutical potential against cancer progression. The molecular mechanisms underlying the response to TAMs-targeted therapies depends on complex dynamics of immune cross-talk and its understanding is still incomplete. In vitro models are helpful to decipher complex responses to combined immunotherapies. In this study, we established and characterized a 3D human macrophage-ER+ PR+ HER2+ breast cancer model, referred to as macrophage-tumor spheroid (MTS). Macrophages integrated within the MTS had a mixed M2/M1 phenotype, abrogated the anti-proliferative effect of trastuzumab on tumor cells, and responded to IFNγ with increased M1-like polarization. The targeted treatment of MTS with a combined CSF1R kinase inhibitor and an activating anti-CD40 antibody increased M2 over M1 phenotype (CD163+/CD86+ and CD206+/CD86+ ratio) in time, abrogated G2/M cell cycle phase transition of cancer cells, promoted the secretion of TNF-α and reduced cancer cell viability. In comparison, combined treatment in a 2D macrophage-cancer cell co-culture model reduced M2 over M1 phenotype and decreased cancer cell viability. Our work shows that this MTS model is responsive to TAMs-targeted therapies, and may be used to study the response of ER+ PR+ HER2+ breast cancer lines to novel TAM-targeting therapies.

Developing an Affinity-Based Chemical Proteomics Method to In Situ Capture Amorphous Aggregated Proteome and Profile Its Heterogeneity in Stressed Cells

Stress induced amorphous proteome aggregation is a hallmark for diseased cells, with the proteomic composition intimately associated with disease pathogenicity. Due to its particularly dynamic, reversible, and dissociable nature, as well as lack of specific recognition anchor, it is difficult to capture aggregated proteins in situ. In this work, we develop a chemical proteomics method (AggLink) to capture amorphous aggregated proteins in live stressed cells and identify the proteomic contents using LC-MS/MS. Our method relies on an affinity-based chemical probe (AggLink 1.0) that is optimized to selectively bind to and covalently label amorphous aggregated proteins in live stressed cells. Especially, chaotrope-compatible ligation enables effective enrichment of labeled aggregated proteins under urea denaturation and dissociation conditions. Compared to conventional fractionation-based method to profile aggregated proteome, our method showed improved enrichment selectivity, detection sensitivity, and identification accuracy. In HeLa cells, the AggLink method reveals the constituent heterogeneity of aggregated proteome induced by inhibition of pro-folding (HSP90) or pro-degradation (proteasome) pathway, which uncovers a synergistic strategy to reduce cancer cell viability. In addition, the unique fluorogenicity of our probe upon labeling aggregated proteome detects its cellular location and morphology. Together, the AggLink method may help to expand our knowledge of the previously nontargetable amorphous aggregated proteome.