Microneedles mediated-dermal delivery of Vitamin C: Formulation, characterization, cytotoxicity, and enhancement of stability.

Vitamin C (VIT C) is an antioxidant that prevents skin aging. Although dermal delivery is one of the most effective routes to transport VIT C to the skin, the impact of this route can be limited by the barrier function of the stratum corneum (SC). Additionally, VIT C rapidly oxidized and degraded under light and temperature. Therefore, this study provides an approach to utilizing microneedles (MNs) to improve the dermal delivery of VIT C and enhance its stability by incorporating a stabilizing system of ethylenediaminetetraacetic acid (EDTA) and sodium metabisulfite (Meta) within the MNs. Vitamin C microneedles (VIT C MNs) were fabricated using different biodegradable polymers and various concentrations of EDTA/Meta. VIT C MNs were evaluated for morphology, VIT C content, mechanical properties, dissolution rate, needles' insertion, physicochemical properties, ex vivo permeation, viscosity of VIT C polymeric solutions, cytotoxicity, and stability. The results showed that VIT C MNs were uniform and mechanically strong. The recovery of VIT C in MNs was 88.3-90.0 %. The dissolution rate of MNs was <30 min. The flux of VIT C varied based on the composition of MNs. VIT C MNs demonstrated safety against human dermal fibroblasts. VIT C MNs with EDTA/Meta (0.1/0.3 %) were stable under different storage conditions for two months. In conclusion, VIT C MNs were successfully developed using biodegradable polymers, and the stabilizing system (EDTA/META) provided a stable dermal delivery system for VIT C to protect skin from aging.

Glutathione and Xanthine Metabolic Changes in Tamoxifen Resistant Breast Cancer Cell Lines are Mediated by Down-Regulation of GSS and XDH and Correlated to Poor Prognosis.

Background: Tamoxifen is commonly used in the treatment of hormonal-positive breast cancer. However, 30%-40% of tumors treated with tamoxifen develop resistance; therefore, an important step to overcome this resistance is to understand the underlying molecular and metabolic mechanisms. In the present work, we used metabolic profiling to determine potential biomarkers of tamoxifen resistance, and gene expression levels of enzymes important to these metabolites and then correlated the expression to the survival of patients receiving tamoxifen. Methods: Tamoxifen-resistant cell lines previously developed and characterized in our laboratory were metabolically profiled with nuclear magnetic resonance spectroscopy (NMR) using cryogenic probe, and the findings were correlated with the expression of genes that encode the key enzymes of the significant metabolites. Moreover, the effect of significantly altered genes on the overall survival of patients was assessed using the Kaplan-Meier plotter web tool. Results: We observed a significant increase in the levels of glutamine, taurine, glutathione, and xanthine, and a significant decrease in the branched-chain amino acids, valine, and isoleucine, as well as glutamate and cysteine in the tamoxifen-resistant cells compared to tamoxifen sensitive cells. Moreover, xanthine dehydrogenase and glutathione synthase gene expression were downregulated, whereas glucose-6-phosphate dehydrogenase was upregulated compared to control. Additionally, increased expression of xanthine dehydrogenase was associated with a better outcome for breast cancer patients. Conclusion: Overall, this study sheds light on metabolic pathways that are dysregulated in tamoxifen-resistant cell lines and the potential role of each of these pathways in the development of resistance.

NMR-based metabolomics identification of potential serum biomarkers of disease progression in patients with multiple sclerosis.

Multiple sclerosis (MS) is a chronic and progressive neurological disorder, characterized by neuroinflammation and demyelination within the central nervous system (CNS). The etiology and the pathogenesis of MS are still unknown. Till now, no satisfactory treatments, diagnostic and prognostic biomarkers are available for MS. Therefore, we aimed to investigate metabolic alterations in patients with MS compared to controls and across MS subtypes. Metabolic profiles of serum samples from patients with MS (n = 90) and healthy control (n = 30) were determined by Nuclear Magnetic Resonance (1H-NMR) Spectroscopy using cryogenic probe. This approach was also utilized to identify significant differences between the metabolite profiles of the MS groups (primary progressive, secondary progressive, and relapsing-remitting) and the healthy controls. Concentrations of nine serum metabolites (adenosine triphosphate (ATP), tryptophan, formate, succinate, glutathione, inosine, histidine, pantothenate, and nicotinamide adenine dinucleotide (NAD)) were significantly higher in patients with MS compared to control. SPMS serum exhibited increased pantothenate and tryptophan than in PPMS. In addition, lysine, myo-inositol, and glutamate exhibited the highest discriminatory power (0.93, 95% CI 0.869-0.981; 0.92, 95% CI 0.859-0.969; 0.91, 95% CI 0.843-0.968 respectively) between healthy control and MS. Using NMR- based metabolomics, we identified a set of metabolites capable of classifying MS patients and controls. These findings confirmed untargeted metabolomics as a useful approach for the discovery of possible novel biomarkers that could aid in the diagnosis of the disease.

Cytotoxic and molecular differences of anticancer agents on 2D and 3D cell culture.

BACKGROUND: Cancer and multidrug resistance are regarded as concerns related to poor health outcomes. It was found that the monolayer of 2D cancer cell cultures lacks many important features compared to Multicellular Tumor Spheroids (MCTS) or 3D cell cultures which instead have the ability to mimic more closely the in vivo tumor microenvironment. This study aimed to produce 3D cell cultures from different cancer cell lines and to examine the cytotoxic activity of anticancer medications on both 2D and 3D systems, as well as to detect alterations in the expression of certain genes levels. METHOD: 3D cell culture was produced using 3D microtissue molds. The cytotoxic activities of colchicine, cisplatin, doxorubicin, and paclitaxel were tested on 2D and 3D cell culture systems obtained from different cell lines (A549, H1299, MCF-7, and DU-145). IC50 values were determined by MTT assay. In addition, gene expression levels of PIK3CA, AKT1, and PTEN were evaluated by qPCR. RESULTS: Similar cytotoxic activities were observed on both 3D and 2D cell cultures, however, higher concentrations of anticancer medications were needed for the 3D system. For instance, paclitaxel showed an IC50 of 6.234 µM and of 13.87 µM on 2D and 3D H1299 cell cultures, respectively. Gene expression of PIK3CA in H1299 cells also showed a higher fold change in 3D cell culture compared to 2D system upon treatment with doxorubicin. CONCLUSION: When compared to 2D cell cultures, the behavior of cells in the 3D system showed to be more resistant to anticancer treatments. Due to their shape, growth pattern, hypoxic core features, interaction between cells, biomarkers synthesis, and resistance to treatment penetration, the MCTS have the advantage of better simulating the in vivo tumor conditions. As a result, it is reasonable to conclude that 3D cell cultures may be a more promising model than the traditional 2D system, offering a better understanding of the in vivo molecular changes in response to different potential treatments and multidrug resistance development.

Investigating Carvedilol's Repurposing for the Treatment of Non-Small Cell Lung Cancer via Aldehyde Dehydrogenase Activity Modulation in the Presence of ß-Adrenergic Agonists.

Repurposing existing drugs appears to be a potential solution for addressing the challenges in the treatment of non-small cell lung cancer (NSCLC). ß-adrenoceptor antagonist drugs (ß-blockers) have tumor-inhibiting effects, making them promising candidates for potential NSCLC treatment. This study investigates the anticancer potential of a subset of ß-blockers in NSCLC cell lines; A549 and H1299. Additionally, it investigates the underlying mechanism behind ß-blockers' anticancer effect by influencing a potential novel target named aldehyde dehydrogenase (ALDH). The MTT assay assessed ß-blockers' cytotoxicity on both cell lines, while Western blot and NADH fluorescence assays evaluated their influence on ALDH protein expression and activity. Carvedilol (CAR) was the most effective blocker in reducing cell survival of A549 and H1299 with IC50 of 18 µM and 13.7 µM, respectively. Significantly, CAR led to a 50% reduction in ALDH expression and 80% decrease in ALDH activity in A549 cells, especially when combined with ß-agonists, in comparison to the control. This effect might be attributed to ß-agonist blockade or an alternative pathway. This novel finding adds to our understanding of CAR's multifaceted anticancer properties, implying that combining CAR with ß-agonists could be a useful strategy for lung cancer treatment.

Synthesis of microwave-assisted carboxamides in Triton WR-1339-induced hyperlipidemic rats: possible hypolipidemic heterocyclic compounds.

The hypolipidemic effect of furan carboxamide derivatives was investigated using the Triton WR-1339 rat model. Nineteen compounds were synthesized, including furan-2-carboxamides of benzophenones and acetophenones (a(1-4)), anilines and amine derivatives (a(5-9)), picolinic-2-carboxamide derivatives of benzophenones and acetophenone (a(10-12)) and furan-2-carboxylate esters of benzophenones and acetophenones, substituted phenols and alcohols (b(1-7)). All the necessary steps were taken to synthesize, purify, and characterize these compounds. They were synthesized by reacting acyl chlorides of the heterocycles with their corresponding amines in the presence of pyridine and tert-butyl acetate. While the conventional heating method yielded acceptable yields for some of the reactions under reflux, the microwave synthesis reactor achieved significantly higher yields for others. Rats with hyperlipidemia were induced with Triton WR-1339 and then subjected to in vivo testing via an intraperitoneal injection of 200 mg kg-1 Triton WR-1339. The model was tested using an oral dose of bezafibrate (100 mg kg-1). After 7 hours of treatment with Triton, the new derivatives represented by compounds a(1-2), a(4-5), a7, and a(10-12) showed significant activity against the complete lipid profile, including a decrease in triglyceride, total cholesterol, and low-density lipoprotein cholesterol and an increase in high-density lipoprotein cholesterol plasma levels. At 20 mg kg-1 dose, these compounds were superior to other lipid-lowering agents in reducing triglyceride levels and slightly increased high-density lipoprotein cholesterol levels. These results indicate a mutual mechanism of action of novel compounds with fibrates, where they have a marked effect on triglyceride and high-density lipoprotein cholesterol levels; for example, a5 causes a significant reduction (p 0.0001) of triglyceride levels by 86%, and a remarkable increase (p 0.0001) in high-density lipoprotein cholesterol plasma levels by 65% as compared to hyperlipidemic rats.

Berberine Inhibited Growth and Migration of Human Colon Cancer Cell Lines by Increasing Phosphatase and Tensin and Inhibiting Aquaporins 1, 3 and 5 Expressions.

Introduction: Berberine is a natural isoquinoline alkaloid with anti-cancer properties. Nevertheless, the underlying mechanism of its action in human colorectal cancer (CRC) has not been thoroughly elucidated. We investigated the anti-cancer effect of berberine on HT-29, SW-480 and HCT-116 human CRC cell lines. Methods: Cell proliferation, migration and invasion were studied by MTT assay, wound healing, transwell chambers and flow cytometry. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and immunostaining were used to evaluate the expression of aquaporins (AQPs) 1, 3 and 5 in colon cancer cell lines before and after treatment with berberine (10, 30 and 100 µM). RT-qPCR and Western blotting were used to further explore the PI3K/AKT signaling pathway and the molecular mechanisms underlying berberine-induced inhibition of cell proliferation. Results: We demonstrated that treatment of these CRC cell lines with berberine inhibited cell proliferation, migration and invasion through induction of apoptosis and necrosis. HT-29, SW-480 and HCT-116 stained positively for AQP 1, 3 and 5, and berberine treatment down-regulated the expression of all three types of AQPs. Berberine also modulated PI3K/AKT pathway activity through up-regulating PTEN and down-regulating PI3K, AKT and p-AKT expression as well as suppressing its downstream targets, mTOR and p-mTOR at the protein level. Discussion/Conclusions: These findings indicate that berberine inhibited growth, migration and invasion of these colon cancer cell lines via down-regulation of AQP 1, 3 and 5 expressions, up-regulating PTEN which inhibited the PI3K/AKT pathway at the gene and protein levels, and that AQP 1, 3 and 5 expression level can be used as prognostic biomarkers for colon cancer metastasis.

The Role of Vitamin E in Protecting against Oxidative Stress, Inflammation, and the Neurotoxic Effects of Acute Paracetamol in Pregnant Female Rats.

Paracetamol (acetaminophen, APAP) is the most common non-prescription analgesic drug used during pregnancy. The aim of this study was to investigate the effect of vitamin E on acute APAP toxicity in pregnant rats. Toxicity in the liver, kidney, and brain (hippocampus, cerebellum, and olfactory bulb) was examined. Twenty pregnant female Wistar rats at gestational day 18 were used. Pregnant rats were divided into four groups: Control, APAP, E + APAP, and APAP + E. The Control group was treated with 0.5 mL p.o. corn oil. The APAP group received 3000 mg/kg p.o. APAP. The E + APAP group received 300 mg/kg p.o. vitamin E one hour before 3000 mg/kg APAP. The APAP + E group received 3000 mg/kg paracetamol one hour before 300 mg/kg p.o. vitamin E. Twenty-four hours after the last treatment administration, rats were euthanized and blood, brain, liver, and kidney samples were collected. Alanine aminotransferase (ALT), aspartate aminotransferase (AST), blood urea nitrogen (BUN), creatinine levels, uric acid (UA), and superoxide dismutase (SOD) levels, as well as the relative mRNA expression of Cyp1a4, Cyp2d6, and Nat2, were determined. Acute APAP treatment upregulated ALT, AST, BUN, and creatinine levels. APAP treatment downregulated UA and SOD levels. APAP treatment upregulated the relative mRNA expression of Cyp1a4 and Cyp2d6, but downregulated Nat2 expression. Vitamin E treatment, either before or after APAP administration, attenuated the toxic effects of APAP. In conclusion, the results showed that an acute toxic APAP dose in late pregnancy can cause oxidative stress and dysregulation in Cyp isoform expression, and that vitamin E treatment attenuates these effects.

Bortezomib advanced mechanisms of action in multiple myeloma, solid and liquid tumors along with its novel therapeutic applications.

Bortezomib (BTZ) is a first-in-class reversible and selective proteasome inhibitor. It inhibits the ubiquitin proteasome pathway that leads to the degradation of many intracellular proteins. Initially, BTZ was FDA approved for the treatment of refractory or relapsed multiple myeloma (MM) in 2003. Later, its usage was approved for patients with previously untreated MM. In 2006, BTZ was approved for the treatment of relapsed or refractory Mantle Cell Lymphoma (MCL) and, in 2014, for previously untreated MCL. BTZ has been extensively studied either alone or in combination with other drugs for the treatment of different liquid tumors especially in MM. However, limited data evaluated the efficacy and safety of using BTZ in patients with solid tumors. In this review, we will discuss the advanced and novel mechanisms of action of BTZ documented in MM, solid tumors and liquid tumors. Moreover, we will shed the light on the newly discovered pharmacological effects of BTZ in other prevalent diseases.

Investigation of the Effect of Imatinib and Hydroxyurea Combination Therapy on Hematological Parameters and Gene Expression in Chronic Myeloid Leukemia (CML) Patients.

(1) Background: Chronic myeloid leukemia is defined as the neoplastic development of mostly myeloid cells in the bone marrow. Several treatments, including chemotherapy, radiation, hormone treatment, and immunological therapy, can be used to control this condition. The therapeutic impact on leukemic individuals varies, and the response to therapy varies between patients due to disease heterogeneity. The primary goal of this study is to compare the effects of single and Imatinib (IM) and Hydroxyurea (HU) combined treatment on hematological parameters and gene expression in CML patients. (2) Methods: This study was conducted on 51 patients, with chronic myeloid leukemia, who were admitted to Al-Basher hospital in Amman, Jordan, for follow-up. Their hematological parameters were checked and gene expression was measured for (BCL2, PP2A, CIP2A, and WT1). (3) Results: The BCL2 gene was found to be less expressed in both IM and (HU + IM) treatments as compared to the HU group alone, while PP2A gene expression was raised. Such a thing indicates that the outcome of the combined therapy method is not ideal, since PP2A activation causes CML cells to move toward the blast crisis stage. Furthermore, CIP2A gene expression revealed that IM and (HU + IM) had the same therapeutic effect and were more successful in CML patients than HU alone. With regards to the treatment effect on hematological parameters, notably in CML patients in later stages, the combination therapy (HU + IM) raised lymphocyte count, indicating a greater response to the treatment. When compared to single medicines, the combination treatment reduced the proportion of neutrophils to normal reference ranges. Platelet counts, on the other hand, dramatically decreased in both IM and (HU + IM). (4) Conclusion: Because the studied genes (BCL2, PP2A, CIP2A, and WT1) are participating in cell proliferation and death, the findings show that the examined genes are significant to understand the efficacy of various therapies. Furthermore, it was found that there was a clear effect of the clinic-based strategic treatment on hematological indicators such as WBCs, lymphocytes, neutrophils, and platelet counts.

Shedding light on pharmacists’ knowledge of kidney stones’ etiology and treatment

Background: The recurring nature of kidney stones (KS) makes it difficult to control and treat. Patients’ education plays a part in reducing disease recurrence. Pharmacists participate in the healthcare services through educating patients with kidney stones about KS preventive measures and medications that greatly reduce the disease frequency and the treatment cost. Insufficient pharmacists’ knowledge may affect the services’ quality and result in misuse of KS medications. Objectives: To evaluate the pharmacists’ level of knowledge to provide adequate information about KS preventive measures, medications, and treatments for patients with kidney stones in Jordan. Methods: An online descriptive survey was distributed to pharmacists to assess their knowledge about KS causes, prevention, and treatment. The results were analyzed using the SPSS software. Results: There were 393 pharmacists participated in this study. Pharmacists demonstrated an overall intermediate level of knowledge about KS. They showed an excellent level of knowledge regarding KS types and etiology, an intermediate level of knowledge about KS preventive measures and treatment, and poor knowledge about home remedies and drugs that promote KS formation. Conclusion: Pharmacists knowledge about KS management through diet and medications need to be improved. This could be through focusing on pharmacists’ training for the effective implementation of knowledge in the clinical practice. Adopting guidelines by pharmacists may reduce the risk of KS recurrence and provide pharmacist-led patient education about KS management in hospitals and community pharmacies. (AU)

Assessment of persistent antimicrobial and anti-biofilm activity of p-HEMA hydrogel loaded with rifampicin and cefixime.

Catheter-associated urinary tract infections (CAUTIs) are nosocomial infections causing more than one million hospital cases annually. The progress of CAUTIs leads to severe health complications. Infections result in blockage of the medical device due to biofilm formation, which necessitates the replacement of the device. The objective of this study is to improve urological biomaterials to minimize microbial growth and reduce the incidence of CAUTIs. Challenges from mixed biofilm are crucial and need to be addressed in the development of new coating materials. Herein, an investigation highlighted the reduction of mixed biofilm overgrowth and attachment tendency on poly-2-hydroxyethyl methacrylate (p-HEMA) surface by loading the hydrogel with rifampicin (RIF), cefixime trihydrate (CFX), and combined ratios of RIF and CFX. Mixed biofilm-formation ability in (3:1) RIF: CFX-loading p-HEMA (F6) surface showed best tendency to resist form biofilm. Persistent antimicrobial activity increased in p-HEMA loaded with combined ratios of RIF and CFX surface compared to p-HEMA alone, antimicrobial activity lasted for 8 days. All fabricated films exhibited %cell viability higher than 75% on HEK 293 cells. The addition of RIF and CFX may improve the duration of urological device employment before replacement.

Extraction of Anthocyanins from Black Grape By-Products and Improving Their Stability Using Cobalt(II) Complexation.

This study was conducted to investigate the effect of cobalt complexation on the spectral properties of anthocyanins (AC) extracted from black grape pomace (Black Magic) and the effect of complexation on the pH stability of AC during storage. Initially, cobalt acetate tetrahydrate aqueous solution was complexed with AC crude extract and diluted separately in buffer solutions with different pH (3.5, 4.5, 5.5, and 6.5). Afterward, spectral changes were determined spectrophotometrically. pH stability was investigated using the same buffer solutions and stored for 7 days in the dark at room temperature, and the absorbance of each solution was measured daily using a spectrophotometer. Results indicated that complexation caused similar hypsochromic and hyperchromic shifts in λmax at all pH values. With regard to pH stability, the degradation of complexed AC followed first-order reaction kinetics causing half-lives to increase up to 80-fold as compared with noncomplexed AC, which was due to the sharp decrease in K (per day), indicating an improved pH stability as compared with noncomplexed AC. Therefore, Co(II) could be used in the stabilization of grape AC for the coloration of a wide range of foods and food products at near-neutral pH environments considering the health benefits of grape AC and the maximum nontoxic dose of Co(II) salt.

Shedding light on pharmacists' knowledge of kidney stones' etiology and treatment.

Background: The recurring nature of kidney stones (KS) makes it difficult to control and treat. Patients' education plays a part in reducing disease recurrence. Pharmacists participate in the healthcare services through educating patients with kidney stones about KS preventive measures and medications that greatly reduce the disease frequency and the treatment cost. Insufficient pharmacists' knowledge may affect the services' quality and result in misuse of KS medications. Objectives: To evaluate the pharmacists' level of knowledge to provide adequate information about KS preventive measures, medications, and treatments for patients with kidney stones in Jordan. Methods: An online descriptive survey was distributed to pharmacists to assess their knowledge about KS causes, prevention, and treatment. The results were analyzed using the SPSS software. Results: There were 393 pharmacists participated in this study. Pharmacists demonstrated an overall intermediate level of knowledge about KS. They showed an excellent level of knowledge regarding KS types and etiology, an intermediate level of knowledge about KS preventive measures and treatment, and poor knowledge about home remedies and drugs that promote KS formation. Conclusion: Pharmacists knowledge about KS management through diet and medications need to be improved. This could be through focusing on pharmacists' training for the effective implementation of knowledge in the clinical practice. Adopting guidelines by pharmacists may reduce the risk of KS recurrence and provide pharmacist-led patient education about KS management in hospitals and community pharmacies.

Lactate and pyruvate levels correlation with lactate dehydrogenase gene expression and glucose consumption in Tamoxifen-resistant MCF-7 cells using capillary electrophoresis with contactless conductivity detection (CE-C4 D).

Breast cancer is the second leading cause of cancer death in women after lung cancer. The first-line treatment of metastatic breast cancer in premenopausal women relies on tamoxifen. The development of tamoxifen resistance is not fully understood. In this study, capillary electrophoresis with capacitively coupled contactless conductivity detector was developed to monitor the changes in lactate and pyruvate levels in supernatant media of three models of developed MCF-7 tamoxifen-resistant cells and correlate these metabolites changes with lactate dehydrogenase genes expression and glucose consumption. The electrophoretic separation was achieved under reversed electroosmotic flow conditions. The linear ranges were 0.15-5 and 0.01-1 mM with a correlation coefficient of 0.9966 and 0.9971 and the limits of detection were 0.01 and 0.02 µM for lactate and pyruvate, respectively. Inter- and intrarun accuracy were in the range of 96.88-105.94% with precision (CV, %) of ≤7.35%. The method was completely validated and the results were in agreement with those obtained using the lactate and glucose assay kits. The results revealed a significant increase in both lactate and pyruvate production in the three tamoxifen-resistant MCF-7 cells models compared to control cells. This increase was correlated with the increase of lactate dehydrogenase genes expression and the increase of glucose consumption.

High Performance Liquid Chromatography-Tandem Mass Spectrometry Method for Correlating the Metabolic Changes of Lactate, Pyruvate and L-Glutamine with Induced Tamoxifen Resistant MCF-7 Cell Line Potential Molecular Changes.

Breast cancer is one of the most prevalent cancers worldwide usually treated with Tamoxifen. Tamoxifen resistance development is the most challenging issue in an initially responsive breast tumor, and mechanisms of resistance are still under investigation. The objective of this study is to develop and validate a selective, sensitive, and simultaneous high performance liquid chromatography-tandem mass spectrometry method to explore the changes in substrates and metabolites in supernatant media of developed Tamoxifen resistance MCF-7 cells. We focus on the determination of lactate, pyruvate, and L-glutamine which enables the tracking of changes in metabolic pathways as a result of the resistance process. Chromatographic separation was achieved within 3.5 min. using a HILIC column (4.6 × 100 mm, 3.5 µm particle size) and mobile phase of 0.05 M acetic acid-ammonium acetate buffer solution pH 3.0: Acetonitrile (40:60 v/v). The linear range was 0.11-2.25, 0.012-0.227, and 0.02-0.20 mM for lactate, pyruvate, and L-glutamine, respectively. Within- and between-run accuracy was in the range 98.94-105.50% with precision (CV, %) of ≤0.86%. The results revealed a significant increase in both lactate and pyruvate production after acquiring the resistant. An increase in L-glutamine levels was also observed and could be attributed to its over production or decline in its consumption. Therefore, further tracking of genes responsible of lactate, pyruvate, and glutamine metabolic pathways should be performed in parallel to provide in-depth explanation of resistance mechanism.

PI3K/AKT and MAPK1 molecular changes preceding matrix metallopeptidases overexpression during tamoxifen-resistance development are correlated to poor prognosis in breast cancer patients.

BACKGROUND: Metastasis and drug resistance remain a persistent key clinical obstacle to the success of breast cancer treatments. Recent years have seen an increased focus on understanding the factors that influence metastasis and drug resistance. METHODS: In this study, the changes in MMPs gene expression were investigated together with their regulatory pathways-PI3K, MAPK and NFKß pathways-during the process of developing tamoxifen resistance in MCF7 cell line. Gene correlation maps and Kaplan-Meier survival plots among all breast cancer patients and patients treated with tamoxifen were evaluated. RESULTS: MMPs gene expression was found to be up regulated in MCF7 cell line treated with tamoxifen during the development of tamoxifen resistance using two approaches. Up-regulation of gene expression of AKT1 and MAPK1 started in cells treated with 10 µM tamoxifen that was followed with up-regulation of other genes in these pathways and MMPs in cells treated with 35 µM tamoxifen. MMPs and genes from PI3K, MAPK and NFKß pathways showed highly significant increase of expression at 50 µM or when cells were treated sequentially six times with 35 µM. Furthermore, increased genes expression was associated with aggressive pattern, clear morphological changes, higher growth rate, increased migration and adhesion potential and tamoxifen insensitivity. Breast cancer distant metastasis-free survival, and survival among tamoxifen treated patients had high expression levels of MAPK1, AKT1, TIMP2, MMP1, and MMP9 showed poor prognosis. CONCLUSION: Early changes of MAPK1, AKT1 gene expression upon tamoxifen treatment could possibly be used as an early marker of resistance and future poor prognosis.

Cytotoxicity and Cellular Death Modality of Surface-Decorated Gold Nanorods against a Panel of Breast Cancer Cell Lines.

Herein, the antiproliferative effect of surface-decorated gold nanorods (GNRs) was investigated against three different breast cancer cell lines. The results indicate that the cell lines exhibited different biological responses and death modalities toward the treatment. The cell lines exhibited similar cellular uptake of the nanoparticles; however, MDA-MB-231 demonstrated the highest cytotoxicity compared to other cell lines upon treatment with GNRs. The expression of the CDH1 gene, which is involved in cell adhesion and metastasis, was dramatically increased in treated MDA-MB-231 cells compared to other cell lines. Early apoptosis and late apoptosis are the dominant cellular death modalities of MDA-MB-231 cells upon treatment with GNRs.

Changes in Lactate Production, Lactate Dehydrogenase Genes Expression and DNA Methylation in Response to Tamoxifen Resistance Development in MCF-7 Cell Line.

Lactate dehydrogenase (LDH) is a key enzyme in the last step of glycolysis, playing a role in the pyruvate-to-lactate reaction. It is associated with the prognosis and metastasis of many cancers, including breast cancer. In this study, we investigated the changes in LDH gene expression and lactate concentrations in the culture media during tamoxifen resistance development in the MCF-7 cell line, and examined LDHB promoter methylation levels. An upregulation of 2.9 times of LDHB gene expression was observed around the IC50 concentration of tamoxifen in treated cells, while fluctuation in LDHA gene expression levels was found. Furthermore, morphological changes in the cell shape accompanied the changes in gene expression. Bisulfate treatment followed by sequencing of the LDHB promoter was performed to track any change in methylation levels; hypomethylation of CpG areas was found, suggesting that gene expression upregulation could be due to methylation level changes. Changes in LDHA and LDHB gene expression were correlated with the increase in lactate concentration in the culture media of treated MCF-7 cells.

Doxorubicin-paclitaxel sequential treatment: insights of DNA methylation and gene expression changes of luminal A and triple negative breast cancer cell lines.

Breast cancer is one of the significant causes of death among women diagnosed with cancer worldwide. Even though several chemotherapy combinations are still the primary treatment of breast cancer, unsuccessful treatments, and poor prognostic outcomes are still being reported. DNA methylation and gene expression changes among two breast cancer cell lines representing luminal A (MCF-7) and triple-negative (MDA-MB-231) cancers were determined after sequential combination treatment of doxorubicin and paclitaxel and analyzed using Ingenuity Pathway Analysis. Promoter methylation changes were seen in different treated MCF-7 cells and accompanied by changes in the gene expression of CCNA1 and PTGS2. In MDA-MB-231 cells, the hypomethylation of ESR1 was not accompanied by an increase in its gene expression in any treated cells. The hypomethylation of GSTP1 and MGMT was accompanied by an increase in gene expression levels in the group treated with doxorubicin only. Also, significant downregulation of several genes like MUC1 and MKI67 in MCF-7 cells treated with doxorubicin showed much lower gene expression (- 37.63, - 10.88 folds) when compared with cells treated with paclitaxel (- 2.47, - 2.05 folds) or the combination treatment (- 18.99, - 2.81 folds), respectively. On the other hand, a synergistic effect on MMP9 gene expression was significantly seen in MDA-MB-231 cells treated with the combination (- 9.99 folds) in comparison with the cells treated with doxorubicin (- 3.62 folds) or paclitaxel (1.75 folds) alone. Chemotherapy combinations do not always augment the molecular changes seen in each drug alone, and these changes could be utilized as treatment response markers.