Development of a novel laboratory photodynamic therapy device: automated multi-mode LED system for optimum well-plate irradiation.

Photodynamic therapy (PDT) is a targeted treatment method that utilizes a photosensitizer (PS) to induce cytotoxicity in malignant and non-malignant tumors. Optimization of PDT requires investigation of the selectivity of PS for the target tissues, irradiating light source, irradiation wavelengths, fluence rate, fluence, illumination mode, and overall treatment plan. In this study, we developed the Multi-mode Automatized Well-plate PDT LED Laboratory Irradiation System (MAWPLIS), an innovative device that automates time-consuming well plate light dosage/PS dose measurement experiment. The careful control of LED current and temperature stabilization in the LED module allowed the system to achieve high optical output stability. The MAWPLIS was designed by integrating a 3-axis moving system and motion controller, a quick-switching LED controller unit equipped with interchangeable LED modules capable of employing multiple wavelengths, and a TEC system. The proposed system achieved high optical output stability (1 mW) within the range of 0-500 mW, high wavelength stability (5 nm) at 635 nm, and high temperature stability (0.2 °C) across all radiation modes. The system's validation involved in vitro analysis using 5-ALA across varying concentrations, incubation periods, light exposures, and wavelengths in HT-29 colon cancer and WI-38 human lung fibroblast cell lines. Specifically, a combination of 405 nm and 635 nm wavelengths was selected to demonstrate enhanced strategies for colon cancer cell eradication and system validation. The MAWPLIS system represents a significant advancement in photodynamic therapy (PDT) research, offering automation and standardization of time-intensive experiments, high stability and precision, and improved PDT efficacy through dual-wavelength integration.

Antibacterial efficacy of silver nanoparticles, sodium hypochlorite, chlorhexidine, and hypochlorous acid on dentinal surfaces infected with Enterococcus faecalis.

The purpose of this study was to evaluate the antibacterial effect of silver nanoparticles (AgNPs) against Enterococcus faecalis and compare it with different irrigation solutions. This study was performed using 64 dentin blocks. E. faecalis suspension was dispensed to each sample and incubated under anaerobic conditions at 37°C throughout 21 days. After the inoculation period, the following solutions were added to each group and kept for 5 min: Group 1, 5.25% sodium hypochlorite (NaOCl); Group 2, 2.5% NaOCl; Group 3, 1% NaOCl; Group 4, 2% chlorhexidine (CHX); Group 5, 200 ppm hypochlorous acid (HOCl); and Group 6, AgNPs. The samples of positive control were treated with sterile saline. Biofilm viability assay was performed using the LIVE/DEAD BacLight Bacterial Viability Kit. Samples were examined using confocal laser scanning microscopy, respectively. There was no significant difference between the 5.25% NaOCl, 2.5% NaOCl, and 1%NaOCl groups (p > .05). However, these groups showed statistically higher antibacterial activity than the 2% CHX, 200 ppm HOCl, and AgNP groups. Also, 2% CHX showed greater percentage of dead cells compared with the AgNP and HOCl groups. While AgNPs group showed lower dead cell rate than all NaOCl groups and 2% CHX, it caused higher dead cells than 200 ppm HOCl group. The 200 ppm HOCl group showed the lowest percentage of dead cells (p < .05) Although the antibacterial effect of AgNPs is not as high as NaOCl and CHX, it has considerable bactericidal activity against E. faecalis and can be improved by further studies. RESEARCH HIGHLIGHTS: New antimicrobial approaches for root canal irrigation. Antimicrobial effect of silver nanoparticles against E. faecalis. Elimination of the biofilm layer for the success of endodontic treatment.

Synergistic effect of a nonsteroidal anti-inflammatory drug in combination with topotecan on small cell lung cancer cells.

BACKGROUND: The topoisomerase I inhibitor topotecan (TPT) is used in the treatment of recurrent small cell lung cancer (SCLC). However, the drug has a limited success rate and causes distress to patients due to its side effects, such as hematologic toxicities, including anemia and thrombocytopenia. Due to these pharmacokinetic limitations and undesirable side effects of chemotherapeutic drugs, the development of combination therapies has gained popularity in SCLC. Meclofenamic acid (MA), a nonsteroidal anti-inflammatory drug, has demonstrated anticancer effects on various types of cancers through different mechanisms. This study aims to investigate the potential synergistic effects of MA and TPT on the small cell lung cancer cell line DMS114. METHODS AND RESULTS: To assess the cytotoxic and apoptotic effects of the combined treatment of MA and TPT, trypan blue exclusion assay, Annexin V, acridine orange/propidium iodide staining, western blot, and cell cycle analysis were conducted. The results demonstrated that the combination of MA and TPT elicited synergistic effects by enhancing toxicity in DMS114 cells (P < 0.01) without causing toxicity in healthy epithelial lung cells MRC5. The strongest synergistic effect was observed when the cells were treated with 60 µM MA and 10 nM TPT for 48 h (CI = 0,751; DRI = 10,871). CONCLUSION: This study, for the first time, furnishes compelling evidence that MA and TPT synergistically reduce cellular proliferation and induce apoptosis in SCLC cells. Combinations of these drugs holds promise as a potential therapeutic strategy to improve efficacy and reduce the side effects associated with TPT.

The regulatory effect of cabazitaxel on epithelial-mesenchymal transition in metastatic prostate cancer.

Introduction: : Epithelial-mesenchymal transition (EMT) is a critical mechanism that promotes cancer cells to metastasis. Therefore, EMT regulation has become an important target in anticancer therapy approaches in recent years. However, in metastatic prostate cancer (PC), the EMT regulatory effect has not fully understood for cabazitaxel (Cbx), a third line taxane-based chemotherapeutic for metastatic castration-resistant PC. Aim: In this study, we investigated the antimetastatic and EMT-regulatory effects of Cbx on hormone-sensitive metastatic PC cells. Materials and Methods: The anticancer effects of Cbx were assessed by WST-1 and Annexin V analysis. The antimetastatic effect of Cbx was evaluated by wound healing and quantitative reverse transcription polymerase chain reaction through EMT-mesenchymal-to-epithelial transition (MET) markers as well as EMT-repressor microRNAs (miRNAs) in Cbx-treated LNCaP cells. Results: Our results showed that, in addition to its apoptotic and anti-migratory activities, Cbx exhibited the EMT-repressor effects through the prominent downregulation of matrix metalloproteinase-9 and Snail levels as EMT-promoting factors, and the significant upregulation of the certain miRNAs, including miR-205, miR-524, and miR-124, which play a role in EMT-repressing by targeting regulators of the EMT-associated genes. Conclusion: Although further evaluations are needed to improve the findings, we showed that, in addition to its classical taxane function, Cbx has a regulatory effect on EMT-MET cycling in hormone-sensitive metastatic PC.

The cytotoxic effects of indoleamine 2, 3-dioxygenase inhibitors on triple negative breast cancer cells upon tumor necrosis factor α stimulation.

Context: Overexpressed indoleamine 2,3-dioxygenase (IDO) has been observed in many types of cancer and plays an essential role in the tumor microenvironment through immune cells function. Aims: In our study, the therapeutic potentials of two different IDO inhibitors (Epacadostat [EPA] and 1-methyl-L-tryptophan [L-1MT]) in triple-negative breast cancer (TNBC) cells were assessed with and without tumor necrosis factor-α (TNF-α) stimulation. Materials and Methods: The anticancer activity of EPA and L-1MT alone and in combination with TNF-α was analyzed by WST-1, annexin V, cell cycle analysis, and acridine orange/ethidium bromide staining. In addition, the relationship between IDO1 and programmed death-ligand 1 (PD-L1) expressions in TNBC cells upon treatment with IDO inhibitors was evaluated by reverse transcription-polymerase chain reaction analysis. Statistical Analysis Used: SPSS 22.0 was conducted for statistical analysis. The one-way analysis of variance with Tukey's multiple comparison test was performed for multiple groups. Independent (unpaired) t -test was used for the comparison of two groups. Results: EPA and L-1MT alone significantly suppressed the TNBC cell viability through the induction of apoptotic cell death and G0/G1 arrest (P < 0.05). TNF-α alone induced the overexpression of IDO1 and PD-L1 in TNBC cells compared with MCF-10A control cells. However, IDO inhibitors significantly inhibited overexpressed IDO1 mRNA levels. Furthermore, EPA alone and co-treated with TNF-α suppressed the mRNA level of PD-L1 in TNBC cells. Therefore, TNF-α stimulation enhanced the therapeutic effects of IDO inhibitors on TNBC. Conclusions: Our findings showed that the efficacy of IDO inhibitors was mediated by pro-inflammatory cytokine. However, different molecular signaling pathways are associated with pro-inflammatory cytokines production, and the expression of IDO1 and PD-L1 calls for further investigations.

Effect of the MiR-99b and MiR-135b on peritoneal carcinomatosis and liver metastasis in colorectal cancer

Abstract Aim This study aimed to evaluate the expression levels of miR-99b and miR-135b in peritoneal carcinoma and liver metastases associated with Colorectal Cancer (CRC), assess their association with the intracellular signaling pathway proteins Kirsten Rat Sarcoma Virus (KRAS) and Akt, and investigate their effects on survival. Materials and methods Changes in the KRAS gene and Akt proteins, expression levels of miR-99b and miR-135b, and factors affecting survival were compared between colorectal cancer-associated peritoneal carcinomatosis and liver metastasis. Results The expression levels of miR-99b and miR-135b and the immunohistochemical grade classification score of Akt were higher in colorectal cancer, peritoneal carcinomatosis, and liver metastasis than in normal tissues (p< 0.05). MiR-99b expression was highest in CRC, whereas miR-135b expression was highest in peritoneal carcinomatosis (p< 0.05). The expression level of miR-99b decreased and that of miR-135b increased in peritoneal and liver metastases compared with that in the tumor tissue. MiR-99b, Akt, and recurrence were risk factors that affected the overall survival rate in the model of clinical predictions (p= 0.045, p= 0.006, and p= 0.012, respectively). Conclusion While the expression of miR-99b was highest in the primary tumor, its decrease in liver metastasis and peritoneal carcinomatosis suggests that miR-99b has a protective effect against liver metastasis and peritoneal carcinomatosis. However, the detection of miR-135b expression was highest in peritoneal carcinomatosis and liver metastasis compared with that in the colorectal cancer tissues suggesting that it facilitates peritoneal carcinomatosis and liver metastasis. Furthermore, miR-99b, KRAS mutations, and Akt are risk factors for the overall survival of colorectal cancer.

Histone deacetylase inhibitor sodium butyrate regulates the activation of toll-like receptor 4/interferon regulatory factor-3 signaling pathways in prostate cancer cells.

CONTEXT: The covalent acetylation and deacetylation of histone proteins by the histone deacetylase (HDAC) enzymes can be considered a novel therapeutic target in prostate cancer (PCa) cells. Sodium butyrate (NaBu) is a HDAC inhibitor (HDACi) which is a promising potential anticancer drug. Toll-like receptor 4 (TLR4) expression is increased in PCa cells and HDACi alter TLR-inducible gene expressions. AIMS: We aimed to evaluate the effects of NaBu on TLR4 mediating signaling pathways in two different PCa cells (DU-145 and LNCaP) for the first time. SUBJECTS AND METHODS: The cytotoxic and apoptotic effects of NaBu were determined by the water-soluble tetrazolium salt (WST-1) and Annexin V-AO/PI assays, respectively. Subcellular localization of TLR4, interferon regulatory factor-3 (IRF3) and Nuclear factor kappa B proteins was evaluated by IF assay. STATISTICAL ANALYSIS USED: All data were statistically analyzed by GraphPad Prism software (V60.1, CA). Obtained data were expressed in a mean ± standard deviation of the three repeated experiments. The differences between control and NaBu treated cells were compared by one-way-ANOVA. P < 0.05 value was considered statistically significant. RESULTS: Our results showed that NaBu significantly inhibited the viability of PCa cells and increased the percentage of apoptotic cells. However, DU-145 cells were more sensitive to NaBu than LNCaP cells. Furthermore, NaBu can induce the cytoplasmic TLR4 and IRF3 expression in particularly DU-145 cells without affecting nuclear translocation of NF-kB in PCa cells. CONCLUSIONS: NaBu induces apoptotic cell death and regulated the TLR4/IRF3 signaling pathways in DU-145 cells but not in LNCaP cells. Therefore, PCa cells differentially responded to NaBu treatment due to probably androgen receptor status.

The Anticancer Potential of Chlorine Dioxide in Small-Cell Lung Cancer Cells.

Background Chlorine dioxide (ClO2) is an effective disinfectant consisting of oxygen, chloride, and potassium. Because of its high oxidative capacity, ClO2 exerts antimicrobial, antiviral, and antifungal effects. However, its anticancer effects remain to be elucidated. Methodology The anticancer activity of CIO2 was assessed on DMS114 small-cell lung cancer (SCLC) cells and human umbilical vein endothelial cells (HUVEC) as control by WST-1, Annexin V, cell cycle analysis, and acridine orange staining. We for the first time investigated the possible therapeutic effects of long-term stabilized ClO2 solution (LTSCD). Results Our preliminary findings showed that LTSCD significantly inhibited the proliferation of SCLC cells (p < 0.01) with less toxicity in HUVEC cells. Additionally, LTSCD induced apoptotic cell death in SCLC cells through nuclear blebbing and vacuolar formation. However, LTSCD treatment did not induce cell cycle arrest in both cell lines. Conclusions LTSCD can be a therapeutic potential for the treatment of SCLC. However, further investigations are required to assess the LTSCD-induced cell death in SCLC both in vitro and in vivo.

The molecular mechanisms of vulpinic acid induced programmed cell death in melanoma.

BACKGROUNDS: Malignant melanoma is an aggressive skin tumor with a rapidly increasing incidence and there is not yet a successful treatment strategy. Vulpinic acid (VA) is derived from secondary metabolites from lichen species. In the current study, we, for the first time, investigated the anti-cancer effects of VA and the underlying mechanism VA induced programmed cell death in melanoma. METHODS: The anti-cancer effects of VA on melanoma cells were evaluated by the xCELLigence system, flow cytometry, caspase-3 activity and RT-PCR analysis. RESULTS: Our results showed that VA had a strong anti-proliferative effect on A-375 melanoma cells without damaging human epidermal melanocyte cells. Additionally, VA promoted apoptotic cell death through G2/M arrest and the activation of both intrinsic and extrinsic apoptosis pathways according to the analysis of 88 genes associated with apoptosis by qRT-PCR. CONCLUSIONS: Our findings suggest that VA could become an alternative topical and transdermal treatment strategy in the treatment of maligned melanoma cancer. However, further investigations are needed to assess the underlying molecular mechanism of VA mediated apoptotic cell death in the treatment of melanoma.

The response of the canine mammary simple carcinoma and carcinosarcoma cells to 5-aminolaevulinic acid-based photodynamic therapy: An in vitro study.

BACKGROUNDS: Canine mammary gland tumors (CMGTs) are heterogeneous tumors and share many similar features with human breast cancer. Despite the improvement of current treatment options, new treatment modalities are required to effectively kill tumor cells without general toxicity in the treatment of CMGTs. Photodynamic therapy (PDT) is a promising method for cancer treatment. However, there is a limited study evaluating the therapeutic efficacy of PDT in the treatment of CMGTs. METHODS: In this context, we, for the first time, investigated the therapeutic potential of 5-aminolaevulinic acid (5-ALA) mediated PDT at 6 and 12 J/cm2 in two different subtypes [Tubulopapillary carcinoma (TPC) and carcinosarcoma (CS)] cells via different molecular analysis. The cytotoxic effects of 5-ALA/PDT on these cells were analyzed by intracellular PpIX level, WST-1 and ROS analysis. Furthermore, the underlying moleculer mechanism of 5-ALA/PDT mediated apoptotic effects on TPC and CS cells were evaluated Annexin V, AO/PI, RT-PCR and western blot analysis. RESULTS: The 5-ALA/PDT treatment upon irradiation considerably inhibited the viability of both TPC and CS cells (p<0.01) and caused apoptotic death through elevated ROS levels, the activation of Caspase-9, and Caspase-3, and the overexpression of Bax. However, the response of TPC and CS cells to 5-ALA/PDT was different. CONCLUSIONS: Our preliminary in vitro findings provide novel insights into the molecular mechanisms underlying 5-ALA/PDT mediated apoptosis in both TPC and CS cells. However, the therapeutic response of CMGT cells to 5-ALA/PDT is limited.

Comparison of the effects of rutaecarpine on molecular subtypes of breast cancer.

OBJECTIVE: Natural compounds have gained considerable attention in recent years due to disadvantages and properties of current chemotherapy drugs in cancer therapy. In addition, the impact of these compounds is specific for each type and/or subtypes of cancer due to different treatment response. Rutaecarpine, an alkaloid obtained from Evodia Rutaecarpa Chinese herb, has anticancer activity by inhibiting topoisomerase and/or cyclo-oxygenase-2 levels. However, the effectiveness of rutaecarpine has not been well known in breast cancer in terms of subtype. Therefore, we investigated the potential therapeutic effects of rutaecarpine on two different subtypes of breast cancer cells. MATERIALS AND METHODS: The cytotoxic and apoptotic effects of rutaecarpine on MCF-7 and MDA-MB-231 cells were analyzed by WST-1, Annexin V, cell cycle, and acridine orange staining. RESULTS: WST-1 results indicated that rutaecarpine significantly inhibited the growth of both cancer cells for 48 h (P < 0.05). In addition, rutaecarpine treatment caused apoptotic cell death through chromatin condensation and nuclear blebbing and G0/G1 arrest in both breast cancer cells. However, the efficacy of rutaecarpine was more profound in MCF-7 cells than MDA-MB-231 cells. CONCLUSIONS: Consequently, rutaecarpine has a potential therapeutic effect on breast cancer. However, the effectiveness of rutaecarpine is dependent on the subtype of breast cancer.

Clinicopathologic features and genetic characteristics of the BRCA1/2 mutation in Turkish breast cancer patients.

The aim of this study was to identify the frequency and spectrum of germline BRCA1/2 pathogenic alterations in a cohort of patients with breast carcinoma. In this study, a total of 603 breast cancer subjects from Turkey were screened for BRCA1/BRCA2 mutations using HDA and Sanger sequencing. In the present study, 21 BRCA1 and BRCA2 pathogenic variants were detected in 30 patients and BRCA1/2 mutations were significantly associated with a family history of breast/ovarian cancer. Analysis of overall survival for BRCA1/BRCA2 mutation carriers showed a trend for poor overall survival only in BRCA1 carriers, although this was not statistically significant in BRCA1 and BRCA2 mutation carriers. The c.5266dupC mutation is one of the most frequently reported mutations in BRCA1 and was identified in five breast cancer patients in our study. The most common BRCA2 gene mutations in the present study were c.8940delA and c.9097dupA, which were found in seven patients. We found mostly BRCA1 and BRCA2 mutation carriers in those patients who showed hormone-positive features. In conclusion, our data showed differences in the distribution of the mutation spectrum of BRCA1 and BRCA2 in Turkey.

Talazoparib Loaded Solid Lipid Nanoparticles: Preparation, Characterization and Evaluation of the Therapeutic Efficacy In vitro.

OBJECTIVE: In the present work, we report for the first time the therapeutic potential of talazoparib (BMN 673)-SLNs for the treatment of BRCA1 deficient Triple Negative Breast Cancer (TNBC). BMN 673-SLNs were produced by hot-homogenization technique and then characterized. METHODS: The cytotoxic and apoptotic effects of BMN 673-SLNs compared with BMN 673 were determined on HCC1937BRCA1-/-, HCC1937-R resistant TNBC and MCF-10A control cell lines. BMN 673- SLNs were found to have reduced particle size (219.5 ± 1.45 nm) and thus more stable (-28.4 ± 2.52 mV) than BMN 673 (1652 ± 2.46 nm and -18.6 ± 0.45 mV) at 4°C. RESULTS: In vitro cell line studies demonstrated that BMN 673-SLNs showed significant cytotoxic effects on HCC1937 (29.8%) and HCC1937-R cells (35.7%) at 10 nM for 12 days compared with BMN 673 (HCC1937 cells: 34.0% and HCC1937-R cells: 93.8% at 10 nM for 12 days) (p<0.05). Additionally, BMN 673-SLNs (40.1%) reduced the toxicity of BMN 673 (53.1%) on MCF-10A control cells thanks to unique physical properties. CONCLUSION: The apoptotic rates in the 10 nM BMN 673-SLNs treatment (88.78% and 85.56%) for 12 days were significantly higher than those in 10 nM BMN 673 (82.6% and 25.86%) for 12 days in HCC1937 and HCC1937-R cells, respectively (p<0.01). Furthermore, these effects were consistent with the findings of colony formation, wound healing and calcein accumulation analysis. In conclusion, the therapeutic potential of BMN 673-SLNs provides a promising chemotherapeutic strategy for the treatment of drugresistant TNBC.

Circulating miR-195 as a Therapeutic Biomarker in Turkish Breast Cancer Patients.

BACKGROUND: Dysregulation of miRNA expression may be used as a biomarker for specific tumours because it may contribute to development of cancer. Circulating miRNA profiles have been highlighted for their potential as predictive markers in heterogeneous diseases such as breast cancer. In the literature, there is evidence that miR-195 levels are differentially expressed pre- and post-operative periods in breast cancer patients. At the same time, miRNA expression levels may vary because of ethnic origins. This study aimed to determine expression levels and potential roles of miR-195 in Turkish breast cancer patients. MATERIALS AND METHODS: The expression patterns of miR-195 were initially examined in breast cancer tissues (luminal A and B type) (n=96). Subsequently, blood samples were prospectively collected from preoperative and postoperative Turkish breast cancer patients and disease free controls. Total RNA was isolated, and the expression level of miR-195 was quantified by real-time PCR. RESULTS: We found that miR-195 level was altered in Turkish breast cancer patients, with down-regulation evident in breast cancer tissues compared to normal adjacent specimens. Furthermore, circulating levels of miR- 195 was significantly decreased in post-operative blood samples compared with pre-operative levels (p=0.01 and <0.05). However, miR-195 was significantly increased in pre-operative blood samples of the luminal B type (p= 0.04 and <0.05). CONCLUSIONS: This study represents the first report of a miR-195 expression profile in Turkish breast cancer patients. Our data suggests that miR-195 levels might be a clinically useful biomarker in the earliest stage of Turkish breast cancer patients.

The Anticancer Activity of Complex [Cu2(µ-(C6H5)2CHCOO)3(bipy)2)] (ClO4) -Solid Lipid Nanoparticles on MCF-7 Cells.

Recent studies have focused on the potential use of metal-based complexes for the treatment of cancer. However, there are some limitations of metal-based ligands for the treatment of cancer due to their toxic effects. In the present study, a novel bimetallic Cu(II) complex, [Cu2(µ-(C6H5)2CHCOO)3 (bipy)2)](ClO4), has firstly been synthesized and characterized by FT-IR, and X-ray crystallography. Furthermore, Cu(II) complex-loaded solid lipid nanoparticles (SLNs) were initially prepared by hot homogenization method to overcome their toxic effects. After characterization, comparative cytotoxic and apoptotic activities of the complex and Cu(II) complex-SLNs on human breast cancer cells (MCF-7) and human umbilical vein endothelial cells (HUVEC) were determined. Cu(II) complex demonstrated considerable in vitro cytotoxic effects on MCF-7 (p<0.05) and induced apoptotic cell death (88.02 ± 3.95%) of MCF-7 cells. But, the complex has also toxic effects (69.5%) on HUVEC control cells. For this purpose, Cu(II) complex-loaded solid lipid nanoparticles (SLN) were firstly produced, with a distrubution range of 190±1.45 nm to 350±1.72 nm and zeta potentials of -27.4±1.98 mV and -18.2±1.07 mV, respectively. The scanning electron microscopy (SEM) images of SLNs were also obtained. In vitro studies have shown that Cu(II) complex-SLNs help in reducing the side effect of Cu(II) complex (29.9%) on HUVEC control cells. Therefore, metal based complex might potentially be used for cancer treatment through nanoparticle based drug delivery systems.