MDA-231 Cells Research Articles

A tumorigenicity evaluation platform for cell therapies based on brain organoids

BackgroundTumorigenicity represents a critical challenge in stem cell-based therapies requiring rigorous monitoring. Conventional approaches for tumorigenicity evaluation are based on animal models and have numerous limitations. Brain organoids, which recapitulate the structural and functional complexity of the human brain, have been widely used in neuroscience research. However, the capacity of brain organoids for tumorigenicity evaluation needs to be further elucidated.MethodsA cerebral organoid model produced from human pluripotent stem cells (hPSCs) was employed. Meanwhile, to enhance the detection sensitivity for potential tumorigenic cells, we created a glioblastoma-like organoid (GBM organoid) model from TP53−/−/PTEN−/− hPSCs to provide a tumor microenvironment for injected cells. Midbrain dopamine (mDA) cells from human embryonic stem cells were utilized as a cell therapy product. mDA cells, hPSCs, mDA cells spiked with hPSCs, and immature mDA cells were then injected into the brain organoids and NOD SCID mice. The injected cells within the brain organoids were characterized, and compared with those injected in vivo to evaluate the capability of the brain organoids for tumorigenicity evaluation. Single-cell RNA sequencing was performed to identify the differential gene expression between the cerebral organoids and the GBM organoids.ResultsBoth cerebral organoids and GBM organoids supported maturation of the injected mDA cells. The hPSCs and immature mDA cells injected in the GBM organoids showed a significantly higher proliferative capacity than those injected in the cerebral organoids and in NOD SCID mice. Furthermore, the spiked hPSCs were detectable in both the cerebral organoids and the GBM organoids. Notably, the GBM organoids demonstrated a superior capacity to enhance proliferation and pluripotency of spiked hPSCs compared to the cerebral organoids and the mouse model. Kyoto Encyclopedia of Genes and Genomes analysis revealed upregulation of tumor-related metabolic pathways and cytokines in the GBM organoids, suggesting that these factors underlie the high detection sensitivity for tumorigenicity evaluation.ConclusionsOur findings suggest that brain organoids could represent a novel and effective platform for evaluating the tumorigenic risk in stem cell-based therapies. Notably, the GBM organoids offer a superior platform that could complement or potentially replace traditional animal-based models for tumorigenicity evaluation.

Exploring the Bioactive Potential of Taraxacum officinale F.H. Wigg Aerial Parts on MDA Breast Cancer Cells: Insights into Phytochemical Composition, Antioxidant Efficacy, and Gelatinase Inhibition within 3D Cellular Models

In this work, aerial parts of Taraxacum officinale F.H. Wigg. produced in Umbria, Italy, were chemically investigated by solid-phase microextraction/gas chromatography–mass spectrometry (SPME/GC-MS) to describe their volatile profile. The results obtained showed the preponderant presence of monoterpenes, with limonene and 1,8-cineole as the main components. Further analyses by GC/MS after derivatization reaction were performed to characterize the non-volatile fraction highlighting the presence of fatty acids and di- and triterpenic compounds. T. officinale methanol and dichloromethane extracts, first analyzed by HRGC/MS, were investigated to evaluate the antioxidant activity, cytotoxicity, and antiproliferative properties of MDA cells on the breast cancer cell line and MCF 10A normal epithelial cells as well as the antioxidant activity by colorimetric assays. The impact on matrix metalloproteinases MMP-9 and MMP-2 was also explored in 3D cell systems to investigate the extracts’ efficacy in reducing cell invasiveness. The extracts tested showed no cytotoxic activity with EC50 > 250 µg/mL on both cell lines. The DPPH assay revealed higher antioxidant activity in the MeOH extract compared with the DCM extract, while the FRAP assay showed a contrasting result, with the DCM extract exhibiting slightly greater antioxidant capacity. After treatment for 24 h with a non-cytotoxic concentration of 500 µg/mL of the tested extracts, gelatin zymography and Western blot analyses demonstrated that both MeOH and DCM extracts influenced the expression of MMP-9 and MMP-2 in MDA cells within the 3D cell model, leading to a significant decrease in the levels of these gelatinases, which are crucial markers of tumor invasiveness.

Enhanced Anticancer and Biological Activities of Environmentally Friendly Ni/Cu-ZnO Solid Solution Nanoparticles

The study investigates the impact of incorporating Ni and Cu into the lattice of ZnO nanoparticles (NPs) to enhance their anticancer and antioxidant properties. Characterization techniques including pXRD, FTIR, UV-visible absorption spectroscopy, FESEM, and EDAX confirm the successful synthesis and structural modifications of Ni/Cu-ZnO NPs. Anticancer activity against breast cancer (MDA) and normal skin (BHK-21) cells reveals dose-dependent cytotoxicity, with Ni/Cu-ZnO NPs exhibiting higher efficacy against MDA cells while being less harmful to BHK-21 cells. Morphological studies corroborate these findings. Additionally, antioxidant assays using TAC, FRAP, and DPPH assay demonstrate the superior antioxidant activity of Ni/Cu-ZnO NPs matched to pure ZnO. Overall, the synergistic effect of Ni and Cu incorporation leads to improved therapeutic potential, making Ni/Cu-ZnO NPs promising candidates for cancer therapy and antioxidant applications.Molecular docking recreations were performed using Auto Dock Vina software to gain more insights and validate the observed biological activities of un-doped ZnO and bi-metal doped ZnO NPs, we investigated the interaction and binding affinities of pure ZnO and bimetallic metal co-doped ZnO for their antioxidant and anticancer studies. Ni/Cu-ZnO have shown good antioxidants and exhibited remarkable anticancer activities.

Design and Synthesis of Pyridyl and 2-Hydroxyphenyl Chalcones with Antitubercular Activity.

A focussed library of pyridyl and 2-hydroxyphenyl chalcones were synthesized and tested for growth inhibitory activity against Mycobacterium tuberculosis H37Rv, and normal and cancer breast cell lines. Pyridyl chalcones bearing lipophilic A-ring, e.g., dichloro-phenyl-(14), pyrene-1-yl (20)- and biphenyl-4-yl (21) moieties were found to be the most potent of the series inhibiting the growth of M. tuberculosis H37Rv with IC90 values ranging from 8.9-28 µM. Aryl chalcones containing a 3-methoxyphenyl A-ring and either p-Br-phenyl (25) or p-Cl-phenyl (26) B-rings showed an IC90 value of 28 µM. Aryl-chalcones were generally less toxic to HepG2 cells compared to pyridyl-chalcones. Dose-dependent antiproliferative activity against MDA468 cells was observed for trimethoxy-phenyl (16) and anthracene-9-yl (19) pyridyl-chalcones with IC50 values of 0.7 and 0.3 µM, respectively. Docking studies revealed that chalone 20 was predicted to bind to the M. tuberculosis protein tyrosine phosphatases B (PtpB) with higher affinity compared to a previously reported PtpB inhibitor.

Phytochemical, biological, and computational investigations of Ephedra alata Decne. growing in salinity conditions of Arabian Peninsula

Ephedra alata Decne is a medicinal plant widely used in traditional medicine for the management of bronchial asthma and cancer. Phytochemical analysis and biological activities, including antioxidant and anticancer effects, were investigated in the current work as new findings for the plant E. alata, a species growing wildly in the marsh and saline environments of the central area of Saudi Arabia. The Ultra Pressure Liquid Chromatography coupled with Electron spray ionization-Quadropole-Time of flight (UPLC-ESI-Q-TOF) system was used for the phytochemical analysis of the plant constituents. In addition, Polyphenolic profiling including the total phenolic (TPC) and flavonoid (TFC) contents of the plant extracts were measured. Phenolic acids were found at the highest relative percentages among all the identified compounds and were measured at 66.07 mg GAE (Gallic acid equivalent). The UPLC analysis of the E. alata extract indicated the presence of chlorogenic acid, syringic acid, caffeic acid, vanillic acid, rosmarinic acid, umbelliferone, isorhoifolin, and apigenin at the highest relative percentages. Mineral analysis indicated that the microelement content of E. alata was relatively low, except for magnesium (Mg). In vitro antioxidant assays revealed the ability of the plant to scavenge DPPH free radicals, reduced molybdenum ions, and ferrous at levels of 14.63, 19.97, and 27.78 mg Trolox equivalents, respectively. The extract induced transition metal chelation at 31.36 mg EDTA equivalents. The extract induced cytotoxic effects against MDA-231 and A549 cell lines at IC50 levels of 25.31 and 39.81 µg/mL, respectively. The plant extract inhibited the colonization and migration of cancer cells as part of its potential anticancer effects. In addition, major E. alata constituents like isorhoifolin, chlorogenic acid, apigenin, and rosmarinic acid exhibited the lowest binding energy to the CAIX enzyme at − 8.41, − 6.64, − 6.32, and − 6.26 kcal/mol, respectively, compared to the binding energy (− 7.72 kcal/mol) of the co-crystallized ligand (Y0R). The docking results further supported the selection of the CAIX enzyme as a standard predictive therapeutic target, since it exhibited significant binding interactions with the major constituents of the plant.

Saponin and fluorine-functionalized polyethyleneimine/covered metal organic framework: A novel gene delivery system for targeted delivery of green fluorescent protein plasmid

Up to now, the existence of a safe and proper nanocarrier to improve the delivery efficiency of plasmid DNA into cells is a challenge. The large surface area and adjustable architectures of metal-organic framework (MOF), have gained a lot of interest and great potential for gene delivery. Herein, the Zirconium-Fumarate MOF (MOF-801) as novel MOF-polymer hybrid nanocarrier with high ability to gene delivery, was synthesized and modified with saponin- and fluorine-functionalized polyethyleneimine through an electrostatic interaction. The size and zeta potential of polyplex (DNA/nanocarrier) was 102 nm and +40 mV, respectively. Not only these powerful nanocarrier transfect the green fluorescent protein (plasmid-non encoded CRISPR/Cas systems) expression plasmid into the cell with an efficiency of about 60 %, but also, they showed good transfection efficiency at low plasmid doses and in existence of 30 % fetal bovine serum. Also, the nanocarrier was applied to study the plasmid delivery into cancer cell lines including MDA, PC12, HCT119, NIHT3, and MCF7 cells. The novel vector displayed low toxicity compared to other vectors which could be utilized for in vivo studies. Therefore, the saponin- and fluorine-modified polyethyleneimine/functionalized MOF-801 nanocomplex with remarkable properties was recommended for efficient gene transference.

Purification and characterization of the produced hyaluronidase by Brucella Intermedia MEFS for antioxidant and anticancer applications

Hyaluronidase (hyase) is an endoglycosidase enzyme that degrades hyaluronic acid (HA) and is mostly known to be found in the extracellular matrix of connective tissues. In the current study, eleven bacteria isolates and one actinomycete were isolated from a roaster comb and screened for hyase production. Seven isolates were positive for hyase, and the most potent isolate was selected based on the diameter of the transparent zone. Based on the morphological, physiological, and 16 S rRNA characteristics, the most potent isolate was identified as Brucella intermedia MEFS with accession number OR794010. The environmental conditions supporting the maximum production of hyase were optimized to be incubation at 30 ºC for 48 h and pH 7, which caused a 1.17-fold increase in hyase production with an activity of 84 U/mL. Hyase was purified using a standard protocol, including precipitation with ammonium sulphate, DEAE as ion exchange chromatography, and size exclusion chromatography using Sephacryle S100, with a specific activity of 9.3-fold compared with the crude enzyme. The results revealed that the molecular weight of hyase was 65 KDa, and the optimum conditions for hyase activity were at pH 7.0 and 37 °C for 30 min. The purified hyase showed potent anticancer activities against colon, lung, skin, and breast cancer cell lines with low toxicity against normal somatic cells. The cell viability of hyase-treated cancer cells was found to be in a dose dependent manner. Hyase also controlled the growth factor-induced cell cycle progression of breast cancer cells and caused relative changes in angiogenesis-related genes as well as suppressed many pro-inflammatory proteins in MDA cells compared with 5-fluorouracil, indicating the significant role of hyase as an anticancer agent. In addition, hyase recorded the highest DPPH scavenging activity of 65.49% and total antioxidant activity of 71.84% at a concentration of 200 µg/mL.

Moringa oleifera seed methanol extract with consolidated antimicrobial, antioxidant, anti-inflammatory, and anticancer activities.

The wide biological activity of the Moringa oleifera represents a potential opportunity for developing selective cancer treatment drugs. The bioactive phytochemicals in Moringa seed extract (MSE) indicated large numbers of phytochemicals (21 compounds) with dominant abundance for cycloisolongifolene, 8,9-dehydro-9-vinyl, and chamazulene accounting for 12.7% and 12.19% of the total detected compounds. The MSE showed a potent anticancer effect toward Caco-2, MDA, and HepG-2 cells with half-maximal inhibitory concentration (IC50) values of 9.15±1.18, 4.85±0.11, and 7.36±0.22µg/mL, respectively, with higher safety (≥31-folds) toward normal human cells (IC50 of 150.7±11.11µg/mL). It appears that MSE stimulates selective-dose-dependent cell shrinkage, and nuclear condensation in the tumor cells, which finally induces the apoptosis pathway to increase its anticancer action. Additionally, MSE showed a potent capability to stimulate cell cycle arrest in both main checkpoint phases (G0/G1 and G2/M) of cell population growth. The apoptotic death stimulation was confirmed through upregulation of tumor protein p53 (p53) and cyclin-dependent kinase inhibitor p21 (p21) expression by more than three- to sixfold and downregulation of B-cell lymphoma 2 expression (threefold) in MSE-treated cells compared to 5-fluorouracil (5-FU)-treated tumor cells. Furthermore, the MSE revealed strong anti-inflammatory activity with significant antioxidant activity by lowering nitric oxide levels and enhancing the superoxide dismutase activity. On the other hand, the MSE revealed broad-spectrum antibacterial activity in a dose-dependent manner against Staphylococcus aureus minimum inhibitory concentration (MIC of 1.25mg/mL), followed by Salmonella typhimurium (MIC of 1.23mg/mL), whereas Escherichia coli was the least sensitive to MSE activity (MIC of 22.5mg/mL) with significant antibiofilm activity against sensitive pathogens.

Evaluation of antimicrobial and anticancer efficacy of silver nanoparticles phytofabricated by Nyctanthes arbor-tristis L. leaf extract

Nyctanthes arbor-tristis L. (Oleaceae), commonly known as harsingar, is a plant with potential medicinal properties. The plant was used in traditional folk medicine and as a pharmacological activity. The present study reports a rapid and eco-accommodating technique to synthesize silver nanoparticles AgNPs with low cost and with no need to heat, using aqueous extracts of N. arbor-tristis L. Phytochemical analysis was conducted to detect the existence of alkaloids, flavonoids, saturated sugar, saponins, glycosides, steroids, tannins, terpenoids, and proteins in the plant. A color change observed in the test confirmed the synthesis of AgNPs. SEM, XRD, EDX, FTIR, and UV – Visible spectrophotometer (DLS) were used to characterize the synthesized AgNPs. The antibacterial potential of AgNPs was further tested against different bacterial strains such as Escherichia coli, Pseudomonas aeruginosa, Salmonella typhimurium, Staphylococcus aureus and Klebsiella pneumonia displayed by finding the maximum zone of inhibition observed against S. typhi of 24.6 mm, S. aureus 21.6±0.57 mm, P. aeruginosa 19 mm, K. pneumonia 17 mm and E. coli 15.3±0.57 mm. Furthermore, the synthesized AgNPs were also exhibited as anticancer properties against MDA-231 cell line Human Breast cancer cell line which was determined dose dependent manner and their concentration of 2.5 to 30 μg/mL respectively, and noted the inhibitory range at 30 μg/mL of its concentration, which was further tested in high inhibitory effect on their leaf extract high concentration in cytotoxic, bactericidal experiments.

SlMDH3 Interacts with Autophagy Receptor Protein SlATI1 and Positively Regulates Tomato Heat Tolerance.

Autophagy, a highly conserved protein degradation system, plays an important role in protecting cells from adverse environmental conditions. ATG8-INTERACTING PROTEIN1 (ATI1) acts as an autophagy receptor, but its functional mechanisms in plants' heat stress tolerance remain unclear. In this study, using LC-MS/MS, we identified malate dehydrogenase (SlMDH3) as a SlATI1-interacting protein. Further studies showed that heat stress induced the expression of SlMDH3 and SlMDH3 co-localized with SlATI1 under both 22 °C and 42 °C heat treatment conditions. Moreover, silencing of SlMDH3 increased the sensitivity of tomato to heat stress, as evidenced by exacerbated degradation of chlorophyll; accumulation of MDA, H2O2, and dead cells; increased relative conductivity; and inhibition of stress-related gene expression. Conversely, overexpression of SlMDH3 improved tomato's heat tolerance, leading to opposite effects on physiological indicators and gene expression compared to SlMDH3 silencing. Taken together, our study suggests that SlMDH3 interacts with SlATI1 and positively regulates tomato heat tolerance.

Polyphenol Contents, Antibacterial and Antioxidant Effects of Four Palestinian Honey Samples, and their Anticancer Effects on Human Breast Cancer Cells

The phenolic compounds of four Palestinian honey samples (PH1-PH4) and their antibacterial effects as well as their cytotoxic, cytostatic, and antimigration effects in human breast cancer cell line (MDA) were evaluated here. HPLC analysis of PH2 (Cornflower), PH3 (Milk thistle), and PH4 (Ziziphus) revealed 15 phenolic compounds, namely, caffeic acid, carvacrol, chrysin, ellagic acid, galangin, gallic acid, kaempferol, p-coumaric acid, pinobanksin, pinocembrin, protocatechuic acid, quercetin, rutin, salicylic acid, and silydamin. The minimum inhibitory concentration (MIC) method applied to Pseudomonas aeruginosa (P. aeruginosa), Staphylococcus aureus (S. aureus), and Escherichia coli (E. coli). A strong positive correlation was detected between antimicrobial activity (E. coli) and p-coumaric acid, quercetin, and silydamin. IC50 values for DPPH neutralization varied from 0.19 w/w% to 10 w/w%. The potential anticancer properties of the honey samples were evaluated on MDA cells. Samples PH2 and PH3 demonstrated cytostatic activity, reducing cell viability by about 43% at non-toxic concentration of 4 mg/mL. The cytostatic effects were strongly correlated with the presence of caffeic acid, chrysin, protocatechuic acid, rutin, and salicylic acid (p < 0.01). Moreover, the cell migration rate was significantly reduced (by up to 85%) with PH2 and PH3 compared to untreated cells (p < 0.05). A strong positive correlation was observed between the cytostatic effects of the concentration of carvacrol and Pinocembrin (p < 0.01). Our findings validate honey’s antibacterial properties and suggest its anticancer benefits may stem from cytostatic and antimigration effects.

Novel Combretastatin A-4 Analogs-Design, Synthesis, and Antiproliferative and Anti-Tubulin Activity.

Combretastatins isolated from the Combretum caffrum tree belong to a group of closely related stilbenes. They are colchicine binding site inhibitors which disrupt the polymerization process of microtubules in tubulins, causing mitotic arrest. In vitro and in vivo studies have proven that some combretastatins exhibit antitumor properties, and among them, combretastatin A-4 is the most active mitotic inhibitor. In this study, a series of novel combretastatin A-4 analogs containing carboxylic acid, ester, and amide moieties were synthesized and their cytotoxic activity against six tumor cell lines was determined using sulforhodamine B assay. For the most cytotoxic compounds (8 and 20), further studies were performed. These compounds were shown to induce G0/G1 cell cycle arrest in MDA and A549 cells, in a concentration-dependent manner. Moreover, in vitro tubulin polymerization assays showed that both compounds are tubulin polymerization enhancers. Additionally, computational analysis of the binding modes and binding energies of the compounds with respect to the key human tubulin isotypes was performed. We have obtained a satisfactory correlation of the binding energies with the IC50 values when weighted averages of the binding energies accounting for the abundance of tubulin isotypes in specific cancer cell lines were computed.

Abstract 2976: The potential use of butyrate and propionate to inhibit oncogenic phenotypes in oral squamous cell carcinoma and triple negative breast cancer

Abstract Background Oral squamous cell carcinoma (OSCC) and triple-negative breast cancer (TNBC) are aggressive cancers with low survival rates, particularly in Hispanics. Since the underpinning mechanisms of oncogenesis for both OSCC and TNBC are multifactorial, the development of novel targeted therapies is limited. The tumor microbiome has been shown to modulate the microenvironment. The microbial product short-chain fatty acids (SCFA) may represent a putative target for treatment as they can potentially inhibit cell growth, proliferation, migration, and induce morphologic changes in cancer cells. We evaluated the impact of the SCFAs butyrate, acetate, and propionate on the oncogenic phenotypes of OECM-1 and MDA-231 cell line models for OSCC and TNBC respectively. Methods OECM-1 and MDA-231 cell lines were cultured in RPMI-1640 medium supplemented with 10% FBS. Cytotoxicity was determined using an alamarBlue assay by treating the cells with SCFA in a series of 7 concentrations starting at 125 mM (1:2 dilution factor) for 24, 48, and 72 hours (h). Proliferation quantification was performed using trypan blue dye after treatment with SCFA for 24h, 48h and 72h. Migration and invasion were measured after treatment for 24h with SCFAs. Comparative analysis of our results was performed using GraphPad Prism v10. Results Higher concentrations of all SCFA decreased cell viability in OSCC and TNBC at all time points. In OECM-1 cells, the average inhibitory concentration of 50% (IC50) was 3mM for butyrate, 382mM for acetate, and 64mM for propionate while in MDA-MB-231 cells, the average IC50 was 4.3mM for butyrate, 462mM for acetate, and 20mM for propionate. Both OECM-1 and MDA-MB-231 cells treated with butyrate and propionate at 5mM resulted in a slower migration after 24h of treatment. Additionally, there was a significant reduction of cell number and proliferation rate after 48h and 72h of butyrate treatment at the concentrations 1mM (p=0.0425) and 5mM (p=0.0007) in OSCC. Conclusions Our results show that butyrate and propionate can reduce oncogenic phenotypes in OSCC and TNBC and may represent a potential target for the development of alternative treatment strategies for both cancers. Future work includes measuring the impact of these SCFA in epithelial-mesenchymal transition, loss of adhesion, and cell death. Citation Format: Angel J. Amaral Maisonet. The potential use of butyrate and propionate to inhibit oncogenic phenotypes in oral squamous cell carcinoma and triple negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2976.

Evaluating the effects of curcumin nano-chitosan on miR-221 and miR-222 expression and Wnt/β-catenin pathways in MCF-7, MDA-MB-231 and SKBR3 cell lines

BackgroundBreast cancer is one of the most common diseases worldwide that affects women of reproductive age. miR-221 and miR-222 are two highly homogeneous microRNAs that play pivotal roles in many cellular processes and regulate the Wnt/β-catenin signaling pathway. Curcumin (CUR), a yellow polyphenolic compound, targets numerous signaling pathways relevant to cancer therapy. The main aim of this study was to compare the ability of chitosan curcumin nanoparticle (CC-CUR) formulation with the curcumin in modulating miR-221 and miR-222 expression through Wnt/β-catenin signaling pathway in MCF-7, MDA-MB-231 and SK-BR-3 breast cancer cell lines.MethodChitosan-cyclodextrin-tripolyphosphate containing curcumin nanoparticles (CC-CUR) were prepared. Cytotoxicity of the CUR and CC-CUR was evaluated. Experimental groups including CC-CUR, CUR and negative control were designed. The expression of miR-221 and miR-222 and Wnt/β-catenin pathway genes was measured.ResultsThe level of miR-221 and miR-222 and β-catenin genes decreased in MCF-7 and MDA-MB-231 cells and WIF1 gene increased in all cells in CC-CUR group. However, the results in SK-BR-3 cell line were unexpected; since miRs and WIF1 gene expressions were increased following CC-CUR administration and β-catenin decreased by administration of CUR.ConclusionAlthough the composite form of curcumin decreased the expression of miR-221 and miR-222 in MCF-7 and MDA cells, with significant decreasing of β-catenin and increasing of WIF1 gene in almost all three cell lines, we can conclude than this formulation exerts its effect mainly through the Wnt/β-catenin pathway. These preliminary findings may pave the way for the use of curcumin nanoparticles in the treatment of some known cancers.

Abstract B039: Butyrate and propionate-reduced oncogenic phenotypes in triple-negative breast cancer

Abstract Objectives Triple negative breast cancer (TNBC) is the most aggressive form of breast cancer affecting 5% of all patients with no known treatment. Hispanic women show a higher risk of developing this type of cancer at an early age. Heterogeneity of TNBC between patients makes it challenging to devolop a effective treatment. The compostion of microbiome amongst the patients has a high variablity that could affect the production of the microbial products. The changes in microbial products have not been previously evaluated and could possibly contribute to chronic inflammation in the patients. Short-chain fatty acids (SCFA), i.e. butyrate, acetate, and propionate which are metabolical products of the microbiome that can potentially inhibit cell growth, proliferation and migration of cancer cells, may represent an alternative treatment for TNBC. Here, we measured the impact of SCFAs treament on viability and migration of MDA-231 cell lines. Methods MDA-231 cell line is an aggressive triple-negative breast cancer (TNBC) cell line common for evaluating new therapeutic strategies. MDA-231 cells were cultured in RPMI-1640 medium supplemented with 10% fetal bovine serum and we evaluated oncogenic phenotypes (cell viability and migration) after SCFA treatment. Cells were treated with butyrate, acetate, and propionate in cell culture media at seven concentrations starting at 125mM (1:2 dilution factor) for 24, 48 and 72 hours (h). We determined the cell viability using Alamar blue and determined the half-maximal inhibitory concentration (IC50) after SCFA treatment at all time points. Furthermore, we performed a wound-healing assay for 24h to evaluate the impact of SCFA on cell migration. MDA-231 cells were cultured to a 100% confluency and a scratch was performed in the cell monolayer to create an artificial gap, simulating the wound. Immediately after performing the wound, the cells were treated with SCFA at concentrations of 1mM and 5mM. Results Higher concentrations of butyrate and propionate reduced cell viability for MDA-231 was reduced. Particularly, butyrate showed an IC50 at 24h, 48h and 72h of 4.7mM, 4.34 mM and 3.6mM, respectively. However, propionate showed an IC50 only at 48h and 72h of 21.09 mM and 19.5 mM, respectively. There was no impact in cell viability after acetate treatment. In relation to migration, MDA-231 cell lines treated with butyrate and propionate at 5mM show a slower migration rate at 24h when compared to the untreated cells. Conclusions Our results suggest that butyrate and propionate can reduce oncogenic phenotypes in MDA-231. Future work will measure the impact of SCFA on cell proliferation, motility, and epithelial-mesenchymal transition. SCFAs may represent a promising target for novel treatment strategies for TNCB. Citation Format: Liah M Román-Calderón, Ariana S García-López, Jeannette L Salgado-Montilla, Gabriel Borges-Vélez, Esther A Peterson-Peguero, Josué Pérez-Santiago. Butyrate and propionate-reduced oncogenic phenotypes in triple-negative breast cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Breast Cancer Research; 2023 Oct 19-22; San Diego, California. Philadelphia (PA): AACR; Cancer Res 2024;84(3 Suppl_1):Abstract nr B039.

NEW ZINC-BONE APATITE FILMS SHOW ANTI-TUMOUR EFFICACY AGAINST BONE METASTASES IN VITRO

Decreasing the chance of local relapse or infection after surgical excision of bone metastases is a main goals in orthopedic oncology. Indeed, bone metastases have high incidence rate (up to 75%) and important cross-relations with infection and bone regeneration. Even in patients with advanced cancer, bone gaps resulting from tumor excision must be filled with bone substitutes. Functionalization of these substitutes with antitumor and antibacterial compounds could constitute a promising approach to overcome infection and tumor at one same time. Here, for the first time, we propose the use of nanostructured zinc-bone apatite coatings having antitumor and antimicrobial efficacy. The coatings are obtained by Ionized Jet Deposition from composite targets of zinc and bovine-derived bone apatite. Antibacterial and antibiofilm efficacy of the coatings is demonstrated in vitro against S. Aureus and E. Coli. Anti-tumor efficacy is investigated against MDA- MB-231 cells and biocompatibility is assessed on L929 and MSCs.A microfluidic based approach is used to select the optimal concentration of zinc to be used to obtain antitumor efficacy and avoid cytotoxicity, exploiting a custom gradient generator microfluidic device, specifically designed for the experiments. Then, coatings capable of releasing the desired amount of active compounds are manufactured. Films morphology, composition and ion-release are studies by FEG- SEM/EDS, XRD and ICP. Efficacy and biocompatibility of the coatings are verified by investigating MDA, MSCs and L929 viability and morphology by Alamar Blue, Live/Dead Assay and FEG-SEM at different timepoints. Statistical analysis is performed by SPSS/PC + Statistics TM 25.0 software, one-way ANOVA and post-hoc Sheffe? test. Data are reported as Mean ± standard Deviation at a significance level of p <0.05.Results and Discussion. Coatings have a nanostructured surface morphology and a composition mimicking the target. They permit sustained zinc release for over 14 days in medium. Thanks to these characteristics, they show high antibacterial ability (inhibition of bacteria viability and adhesion to substrate) against both the gram + and gram – strain.The gradient generator microfluidic device permits a fine selection of the concentration of zinc to be used, with many potential perspectives for the design of biomaterials. For the first time, we show that zinc and zinc-based coatings have a selective efficacy against MDA cells. Upon mixing with bone apatite, the efficacy is maintained and cytotoxicity is avoided. For the first time, new antibacterial metal-based films are proposed for addressing bone metastases and infection at one same time. At the same time, a new approach is proposed for the design of the coatings, based on a microfluidic approach. We demonstrated the efficacy of Zn against the MDA-MB-231 cells, characterized for their ability to form bone metastases in vivo, and the possibility to use nanostructured metallic coatings against bone tumors. At the same time, we show that the gradient-generator approach is promising for the design of antitumor biomaterials. Efficacy of Zn films must be verified in vivo, but the dual-efficacy coatings appear promising for orthopedic applications.

Folic acid-conjugated magnetic-luminescent nanocomposites from Mn0.8Fe2.2O4 and GdVO4:Dy3+ with efficient heat generation and cytocompatibility in MDA-231 cell lines

Folic acid-conjugated magnetic-luminescent nanocomposites from Mn0.8Fe2.2O4 and GdVO4:Dy3+ with efficient heat generation and cytocompatibility in MDA-231 cell lines

Cytotoxic Activity of Hypericum triquetrifolium Turra Methanolic Extract Against Cancer Cell Lines.

Since ancient times, several people in the Mediterranean region have employed Hypericum triquetrifolium Turra in traditional medicine. However, only the composition of its essential oils has received extensive research. This study investigated the cytogenetic and cytotoxic effects of H. triquetrifolium methanolic leaf extract on four different cancer cell lines. Methanolic extract of H. triquetrifolium leaves was prepared. Albino male mice were grouped into five: group 1 (blank control) received water;group II (CYP) received cyclophosphoamide; groups III, IV, and V were administered 50, 100, and 200 mg/kg of H. triquetrifolium extracts. On the 11th day, the animals were sacrificed and bone marrowwascollected. The metaphase index (MI) of the bone marrow cells was determined to evaluate the cytogenetic effect of the extract. The cytotoxic activity of the extract was tested on four cancer cell lines (HepG2, PC3, MDA, and A594), while WRL-68 normal cells were employed as control. The index of bone marrow cells in cyclophosphamide (CYP)-treated albino male mice shows a significant difference (P ≤ 0.05) between concentration inhibition 50 % IC50 of cancer cell line compered to WRL-68 normal cells, on MDA and WRL-68 cells (IC50 =185.7, 200.7), HepG2 and WRL-68 (IC50 104.9, 564.6), A549 and WRL-68 (IC50 115, 192), PC3 and WRL-68 (IC50 160.7, 298.7). The results showed that the extracts was able to increase metaphase index compared to cyclophasoamide-treated mice, which caused a drop in the percentage of metaphase index. The current study was showed that significant anti - proliferation activity of Hypericum triquetrifolium methanolic extract against MDA-MB-231 cell line is an epithelial, human breast cancer cell line,WRL-68, HepG2 and lung cancer cell lines (A594).

Selective labeling of phosphatidylserine for cryo-TEM by a two-step immunogold method

Immunogold labeling in transmission electron microscopy (TEM) utilizes the high electron density of gold nanoparticles conjugated to proteins to identify specific antigens in biological samples. In this work we applied the concept of immunogold labeling for the labeling of negatively charged phospholipids, namely phosphatidylserine, by a simple protocol, performed entirely in the liquid-phase, from which cryo-TEM specimens can be directly prepared. Labeling included a two-step process using biotinylated annexin-V and gold-conjugated streptavidin. We initially applied it on liposomal systems, demonstrating its specificity and selectivity, differentiating between 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) and 1,2-dioleoyl-sn-glycero-3-phospho-l-serine (DOPS) membranes. We also observed specific labeling on extracellular vesicle samples isolated from THP1 cells and from MDA-468 cells, which underwent stimulations. Finally, we compared the levels of annexin-V labeling on the cells vs. on their isolated EVs by flow cytometry and found a good correlation with the cryo-TEM results. This simple, yet effective labeling technique makes it possible to differentiate between negatively charged and non-negatively charged membranes, thus shillucidating their possible EV shedding mechanism.

Investigating the Toxicity Caused by Photodynamic Therapy with a Low-power Diode Laser with a Wavelength of 810 nm Compared to a Combination of Curcumin in Breast Cancer Cells

Background: Curcumin has low side effects and strong antioxidant, anti-diabetic and anti-cancer activities, and other therapeutic properties. Studies have confirmed that visible light can improve curcumin’s antibacterial, anti-inflammatory, antioxidant, and anti-cancer properties. Objectives: This study aimed to investigate the toxicity caused by photodynamic therapy with a low-power diode laser with a wavelength of 810 nm compared to curcumin in breast cancer cells (MDA-MB-231 cell line). Methods: After culturing cells in a 96-well plate, concentrations of 1, 10, 100, and 500 μM of curcumin alone and also in combination with a low-power diode laser with a wavelength of 810 nm and after 48 hours of incubation, cells’ viability was assessed by MTT method. Also, cell proliferation rate, lactate dehydrogenase enzyme release, and membrane lipid peroxidation were measured. Results: The percentage of survival, cell proliferation rate, and release of lactate dehydrogenase enzyme in the combination of 500 μg/mL with laser was significantly lower than the rest of the groups, while the average concentration of malondialdehyde obtained according to micrograms in the curcumin group with laser was higher than other groups. Conclusions: Overall, combining curcumin with a photodynamic laser may reduce the survival effect and cell proliferation rate of curcumin on MDA cells. Most current research on curcumin combined with photodynamic lasers focuses on in vitro experiments and animal models; however, further clinical research is needed.