From Androgen Dependence to Independence in Prostate Cancer: Unraveling Therapeutic Potential and Proteomic Landscape of Hydroxychloroquine as an Autophagy Inhibitor.

Prostate cancer is a major planetary health challenge wherein new ways of thinking drug discovery and therapeutics innovation are much needed. Numerous studies have shown that autophagy inhibition holds a significant role as an adjunctive intervention in prostate cancer. Hydroxychloroquine (HCQ) has gained considerable attention due to its established role as an autophagy inhibitor across diverse cancer types, but its proteomics landscape and systems biology in prostate cancer are currently lacking in the literature. This study reports the proteomic responses to HCQ in prostate cancer cells, namely, androgen-dependent LNCaP and androgen-independent PC3 cells. Differentially expressed proteins and proteome in HCQ-treated cells were determined by label-free quantification with nano-high-performance liquid chromatography and tandem mass spectrometry (nHPLC-MS/MS), and harnessing bioinformatics tools. In PC3 cells, there was a marked shift toward metabolic reprogramming, highlighted by an upregulation of mitochondrial proteins in oxidative phosphorylation and tricarboxylic acid cycle, suggesting an adaptive mechanism to maintain energy production under therapeutic stress. In contrast, LNCaP cells prioritized proteostasis and cell cycle regulation, indicating a more conservative adaptation strategy. To the best of our knowledge, this study is the first to demonstrate the differential responses of prostate cancer cells to autophagy inhibition by HCQ, suggesting that a combination therapy approach, targeting distinct pathways in androgen-independent and androgen-dependent cells, could represent a promising treatment strategy. Moreover, the varied proteomic responses observed between these cell lines underscore the importance of personalized medicine in cancer therapy. Future translational and clinical research on HCQ and prostate cancer are called for.

GFP Transfection Alters Protein Expression Patterns in Prostate Cancer Cells: A Proteomic Study.

PURPOSE: Green Fluorescent Protein is widely used as a cellular marker tool, but its potential influence on cells has been questioned. Although the potential off-target effects of GFP on tumor cells have been studied to some extent, the findings at the molecular level are insufficient to explain the effect of GFP expression on the tumorigenic capacity of cancer cells. Here, we aimed to investigate the effect of GFP expression on the tumorigenicity of PC3 prostate cancer cells. METHODS: Using GFP-expressing and wild-type PC-3 cells, xenograft models were generated in athymic BALB/C mice. To identify differentially expressed proteins, the change in cells proteome was investigated by label-free quantification with nano-high performance liquid chromatography to tandem mass spectrometry (nHPLC-MS/MS). Proteins that showed significantly altered expression levels were evaluated using the bioinformatics tools. RESULTS: Unlike the wild-type PC-3 cells, GFP-expressing cells failed to develop tumor. Comparative proteome analysis of GFP-expressing cells with WT PC-3 cells revealed a total of 216 differentially regulated proteins, of which 98 were upregulated and 117 were downregulated. CONCLUSION: Upon GFP expression, differential changes in several pathways including the immune system, translational machinery, energy metabolism, elements of cytoskeletal and VEGF signaling pathway were observed. Therefore, care should be taken into account to prevent reporting deceitful mechanisms generated from studies utilizing GFP.

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.

Investigation of the combined cytotoxicity induced by sodium butyrate and a flavonoid quercetin treatment on MCF-7 breast cancer cells.

Quercetin (QUE) belonging to the flavonoid class is a common phytochemical present in the daily diet of some individuals. Quercetin is an important source of free radical scavengers. This property makes this flavonoid a reliable antioxidant with the following properties: anti-inflammatory, anti-diabetic, antimicrobial and anti-carcinogenic. Sodium butyrate (NaBu) acts as a histone deacetylase inhibitor (HDACi) and is known to regulate apoptosis in cancer cells. Combining natural flavonoids such as QUE with different substances may synergistically enhance their anti-carcinogenic capacity. Thus, the aim of this study was to examine the combined treatment effects of QUE and NaBu in hormone-sensitive breast cancer cells in vitro. MCF-7 breast cancer cells were treated with QUE alone, NaBu alone, as well as QUE and NaBu combined to determine the following: cell proliferation, levels of protein annexin A5 (ANXA5) and reactive oxygen species (ROS), mRNA protein expression, as well as cell and nuclear morphology. Data demonstrated that either QUE or NaBu alone inhibited cell proliferation, and reduced levels protein ANXA5, ROS and mRNA protein expression, The combination of QUE and NaBu produced a significant synergistic inhibitory effect compared to treatment groups of QUE or NaBu alone. In conclusion, our findings showed that the combination treatment of QUE and NaBu may constitute a promising therapeutic approach to breast cancer treatment but this needs further molecular and in vivo investigations.

Proteomics analysis of meclofenamic acid-treated small cell lung carcinoma cells revealed changes in cellular energy metabolism for cancer cell survival.

Small cell lung carcinoma (SCLC) is a highly aggressive cancer with low survival rate. Although initial response to chemotherapy in SCLC patients is well-rated, the treatments applied after the disease relapses are not successful. Drug resistance is accepted to be one of the main reasons for this failure. Therefore, there is an urgent need for new treatment strategies for SCLC. Meclofenamic acid, a nonsteroidal anti-inflammatory drug, has been shown to have anticancer effects on various types of cancers via different mechanisms. The aim of this study was to investigate the alterations that meclofenamic acid caused on a SCLC cell line, DMS114 using the tools of proteomics namely two-dimensional gel electrophoresis coupled to MALDI-TOF/TOF and nHPLC coupled to LC-MS/MS. Among the proteins identified by both methods, those showing significantly altered expression levels were evaluated using bioinformatics databases, PANTHER and STRING. The key altered metabolism upon meclofenamic acid treatment appeared to the cellular energy metabolism. Glycolysis was suppressed, whereas mitochondrial activity and oxidative phosphorylation were boosted. The cells underwent metabolic reprogramming to adapt into their new environment for survival. Metabolic reprogramming is known to cause drug resistance in several cancer types including SCLC. The identified differentially regulated proteins in here associated with energy metabolism hold value as the potential targets to overcome drug resistance in SCLC treatment.

Investigation of the effect of meclofenamic acid on the proteome of LNCaP cells reveals changes in alternative polyadenylation and splicing machinery.

Prostate cancer is the most common type of cancer among men, and there is still no definitively effective drug treatment. Thus, the search for novel drug agents that may be used for the effective treatment continues. Meclofenamic acid (MA), a non-steroidal anti-inflammatory drug, with anti-tumor effects in various types of cancers was used to investigate its effects on LNCaP cells, a prostate cancer cell line, at the proteome level. The cells were treated with 80 µM MA for 24 h and a comparative proteomic analysis was performed with their untreated control cells. Proteins were extracted from the cells and then were subjected to two-dimensional gel electrophoresis. Protein spots displaying changes in their regulation ratios for more than two-fold were excised from the gels and identified with MALDI-TOF/TOF mass spectrometry. Bioinformatics analysis of the differentially regulated proteins that we identified showed that they were all associated with and took part in related pathways. Glycolytic pathway, cytoskeletal formation, transport activity, protein metabolism, and most notably an mRNA processing pathway were affected by the MA treatment. In addition to presenting a detailed information for what is happening inside the cells upon MA treatment, the proteins affected by MA treatment hold the potential to be novel targets for prostate cancer treatment provided that further in vivo experiments are carried out.

Proteomic analysis of talazoparib resistance in triple-negative breast cancer cells.

Talazoparib (TAL) has been effectively used for the treatment of gBRCA1/2-mutated HER2-negative metastatic breast cancer. However, acquired resistance to TAL remains a major challenge that impedes the clinical success of TAL treatment. Therefore, elucidation of proteins and pathways that contribute to or are affected by the TAL resistance is urgently needed to improve the treatment response and provide novel treatment strategies for advanced metastatic breast cancers. Herein, we aimed to investigate the altered protein signatures in TAL-resistant triple-negative breast cancer (TNBC) cells by comparing with the TNBC parental cell line via proteomic analysis. After validation of TAL-resistance by WST-1 and Annexin V analysis, two-dimensional gel electrophoresis (2DE)-based proteomic analysis coupled to matrix-assisted laser desorption/ionization (MALDI)-time of flight (TOF) mass spectrometry was performed to identify differentially regulated proteins. The findings revealed the identities of 10 differentially regulated proteins in TAL-resistant TNBC cells whose bioinformatic analysis predicted changes in EGF/FGF signaling pathways as well as in the AMPK signaling pathway. In addition, phosphorylation/dephosphorylation dynamics were predicted to be altered in TAL-resistant cells. The proteins identified in this study might be the targets to overcome TAL resistance for the treatment of TNBC.

Histological Study of The Healing of Traumatic Tympanic Membrane Perforation After Vivosorb and Epifilm Application.

BACKGROUND: Untreated traumatic tympanic membrane perforations (TMPs) may lead to permanent perforations and hearing loss. There are many materials that have been previously used for repairing the TMPs. AIMS AND OBJECTIVES: The purpose of this study is to evaluate the clinical and histological effects of Vivosorb (Vv) and Epifilm on healing of TMPs in a rat model. MATERIAL AND METHODS: The posterior-inferior quadrant of the tympanic membranes (TMs) in right ears of 14 rats was perforated using a 20-g needle and then the animals were randomly divided into 2 equal groups (n = 7). The perforated right TMs were treated with either Vv (Vv group) or Epifilm (Ep group). The left TMs of 7 rats were perforated in same way and allowed to close spontaneously without any topical material applications (spontaneous closure group as sham control, SC). The left tympanic membranes of the other 7 rats were not perforated and used as normal controls (NC group). On postoperative 15th day, tympanic bullas were extracted from killed rats and examined morphometrically and histopathologically. RESULTS: Perforation closure rate was 85.7% (6/7) in both Vv and SC groups. Perforations of Ep group closed in 7/7 (100%) ears. The thicknesses of the perforated membranes were increased in SC and especially Vv groups. Also, connective tissue fibrosis, blood clots, and epithelial degenerations were detected in SC and Vv groups. The mean fibroblastic reaction scores of Vv, Ep, and SC groups were 2.14(+), 0.57(+), and 1.71(+) respectively, on comparison with NC group. The mean neovascularization score was 1.42(+) in Vv group, 0.14(+) in Ep group, and 0.57(+) in SC group. CONCLUSION AND SIGNIFICANCE: Vivosorb and especially Epifilm can improve the healing process in traumatic TMPs and additionally, Epifilm might be more preferred for the treatment of TMPs because of causing lesser fibrosis.

The comparison of the viability of crushed, morselized and diced cartilage grafts: a confocal microscopic study.

To compare the cellular viability of diced, crushed, and morselized cartilage used in nasal surgeries. In this study, cartilage was extracted from the ears of seven New Zealand rabbits and was subsequently either diced, crushed or morselized to an amorphous state, or left unmodified. The four types of grafts were then implanted in the back regions of the rabbits. After 3 months, the cellular viability from four groups was compared to a control group using confocal microscopy. Analysis of the data obtained from the enumeration of live cells showed no statistically significant difference between the unmodified graft group and the control group. The diced, crushed, and morselized cartilage groups did show a statistically significant difference in terms of live cell count with the highest number of live cells in diced cartilage group. A statistically significant decrease in live cell count was detected in crushed cartilage group. Our study shows that the viability of cells in diced cartilage grafts is greater than those in crushed or morselized cartilage grafts.