Advances in targeting cancer epigenetics using CRISPR-dCas9 technology: A comprehensive review and future prospects.

Cancer, a complex and multifaceted group of diseases, continues to challenge the boundaries of medical science and healthcare. Its relentless impact on global health, both in terms of prevalence and mortality, underscores the urgent need for a comprehensive understanding of its underlying mechanisms and innovative therapeutic approaches. In recent years, significant progress has been achieved in identifying the genetic and epigenetic mechanisms that cause cancer development and treatment resistance. Researchers are currently investigating the possibility of epigenetic editing such as CRISPR-dCas9 (Clustered Regularly Interspaced Short Palindromic Repeats/deactivated CRISPR-associated protein 9) technologies, for targeting and modifying cancer related epigenetic alterations. A revolutionary form of precision cancer treatment called CRISPR-dCas9 is derived from the bacterial CRISPR-Cas (CRISPR-associated nuclease) system. CRISPR-dCas9 can be combined with epigenetic effectors (EE) to alter malignant epigenetic characteristics associated with cancer. The purpose of this review article is to provide a thorough analysis of recent advancements in utilizing CRISPR-dCas9 technology to target and modify epigenetic changes associated with cancer. This review aims to summarize the latest research developments, evaluate the effectiveness and limitations of CRISPR-dCas9 applications in cancer therapy, identify key challenges such as delivery methods and explore future directions for improving and expanding these technologies. Here, we address the various obstacles that may arise in clinical applications while showcasing the latest advancements and potential future uses of CRISPR-Cas9 in cancer therapy.

Evaluation of Cytotoxic and Anti-cancer Potential of Capsaicin on Lung Cancer Cell Line: An In Vitro Investigation.

Background The leading cause of cancer-related deaths worldwide is lung cancer. Approximately 1.8 million new cases were diagnosed, and 1.6 million individuals died. Available treatment options are inefficient leading to tumour recurrence. Hence there is a need for novel therapeutic advancements in lung cancer treatment. Capsaicin, a naturally occurring protoalkaloid, was found to possess several potential benefits. Aim The aim of the study was to examine capsaicin's cytotoxic and anti-cancer effects in the lung cancer cell line (A549). Materials and methods The cell viability of lung cancer cells treated with capsaicin was measured using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. A549 cells were treated with capsaicin at concentrations ranging from 25 to 150 µM/mL for 24 hours. Changes in cell morphology were observed using a phase-contrast microscope. Nuclear morphological alterations in the lung cancer cells were examined through acridine orange/ethidium bromide (AO/EtBr) staining and viewed under a fluorescent microscope to identify apoptotic nuclei. Gene expression analysis was performed using quantitative real-time PCR (Polymerase Chain Reaction) to evaluate the expression of apoptotic genes, transforming growth factor-beta (TGF-ß), and suppressor of mothers against decapentaplegic 2 (SMAD2). Capsaicin's anti-migratory properties were assessed using a scratch wound healing assay. Result Our study demonstrated that treating lung cancer cells with capsaicin dramatically decreased their vitality, with a statistically significant difference (p<0.05) between the treatment and control groups. In lung cancer cells, we measured the inhibitory concentration (IC-50) at 101.2µM/ml. Following treatment, the number of cells decreased, and those that remained exhibited cytoplasmic membrane blebbing and shrunk. With AO/EtBr staining, treated cells showed an increased number of apoptotic cells. The study's findings showed that after receiving capsaicin, there was a significant downregulation of TGF-ß and SMAD2. Moreover, when compared to control cells, capsaicin-treated cells' migration was markedly reduced. Through modification of the TGF-ß/SMAD2 signaling system, capsaicin therapy dramatically promotes apoptosis and inhibits migration. Conclusion In conclusion, the study's results indicate that capsaicin may have anti-tumor effects on lung cancer cells. To fully comprehend the mechanism underlying capsaicin's anticancer potential and its therapeutic application, further studies are much needed.

Pycnogenol's Dual Impact: Inducing Apoptosis and Suppressing Migration via Vascular Endothelial Growth Factor/Fibroblast Growth Factor Signaling Pathways in Breast Cancer Cells.

BACKGROUND: The leading cause of cancer-related fatalities in women globally is breast cancer. Chemotherapy is one of the traditional therapies for breast cancer, even though it does not target cancer cells directly and has major side effects. As a result, the development of novel therapeutic techniques with improved safety and effectiveness is constantly required. AIM:  This study aimed to investigate the pro-apoptotic and anti-migrative effects of pycnogenol in a breast cancer cell line. METHODOLOGY:  By using the 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) method, the cell viability of breast cancer cells treated with pycnogenol was evaluated. Pycnogenol was applied to the MCF-7 cells in a range of concentrations (20-120 µg/ml) for 24 hours. A phase contrast microscope is used to evaluate changes in cell morphology. In breast cancer cells, acridine orange (AO) and ethidium bromide (EtBr) dual staining were employed to analyze the nuclear morphological alterations. A fluorescent microscope was used to see the apoptotic nuclei. A scratch wound healing assay was performed to evaluate the anti-migrative potential of pycnogenol. Gene expression analysis was performed using quantitative real-time PCR to determine the levels of proapoptotic and vascular endothelial growth factor (VEGF)/VEGF receptor (VEGFR) genes mRNA expression.  Results: In our investigation, breast cancer cells treated with pycnogenol displayed a substantial reduction in cell viability and a statistically significant p<0.05 between the control and treatment groups. We observed inhibitory concentrations (IC-50) at 80 µg/mL in breast cancer cells. After treatment, fewer cells were present, and those that were there shrank and showed cytoplasmic membrane blebbing. Under AO/EtBr staining, treated cells show chromatin condensation and nuclear fragmentation. The results of this study revealed a significant downregulation of Bcl-2, VEGF/FGF, and p53 mRNA expression following treatment with pycnogenol. Furthermore, the impact of pycnogenol on cell migration decreased significantly when compared to control cells. Pycnogenol treatment significantly induces apoptosis and inhibits migration by altering the VEGF signaling pathway.  Conclusion: Overall, this study highlights the promising role of pycnogenol as a proapoptotic and antimigrative agent through the inhibition of anti-apoptotic and VEGF/FGF signaling molecules gene expression, offering new prospects for improving breast cancer treatment.

Exploring the Cytotoxic and Anticancer Potential of Naringin on Oral Cancer Cell Line.

Introduction Oral cancer is the most persistent, aggressive primary malignant sarcoma that is globally prevalent. Though chemotherapy is the only treatment option, it has not progressed for years to overcome its detrimental side effects. Introducing novel therapeutic techniques to improve effectiveness is the need of the hour. Aim This study aimed to investigate the pro-apoptotic effects of naringin in oral cancer cell lines. Methodology The cell viability of oral cancer cells treated with naringin was measured using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method. Naringin was given to oral cancer cells (KB-1) in concentrations ranging from 20 to 200 µM/mL for 24 hours. A phase-contrast microscope is used to examine cell morphology changes. Ethidium bromide (EtBr) staining was employed to study nuclear morphological alterations in oral cancer cells. The apoptotic nuclei were viewed under a fluorescent microscope. To determine pro-apoptotic levels, quantitative real-time polymerase chain reaction (PCR) gene expression analysis was performed to evaluate the expression of transforming growth factor-beta (TGF-ß), suppressor of mothers against decapentaplegic 2 (SMAD2), tumor necrosis factor alpha (TNFα), and nuclear factor kappa B (NFκB). A scratch wound healing experiment was used to evaluate naringin's anti-migratory properties. Results Our study found that naringin treatment significantly reduced cell viability in oral cancer cells compared to the control group (p < 0.05). In oral cancer cells, we found an inhibitory concentration (IC50) of 125.3 µM/mL. Following treatment, fewer cells were present, and those that were present shrunk and displayed cytoplasmic membrane blebbing. The EtBr staining reveals chromatin condensation and nuclear breakage in treated cells. The study found that naringin downregulates the expression of B-cell leukemia/lymphoma 2 (Bcl-2), TGF-ß, SMAD2, TNFα, and NFκB and upregulates the expression of Bcl-2-associated agonist of cell death (BAD), Bcl-2-associated protein X (BAX), and caspase-3. Furthermore, when compared to control cells, naringin significantly reduced cell migration. Naringin treatment significantly promotes apoptosis and inhibits migration by altering the SMAD2 signaling pathway. Conclusion Overall, this study highlights the promising role of naringin as a pro-apoptotic and cytotoxic phytochemical regulating the gene expression of Bcl-2, TGF-ß, SMAD2, TNFα, NFκB, BAD, BAX, and caspase-3, thereby treating oral cancer.

Enhanced Apoptotic Effects in MDA-MB-231 Triple-Negative Breast Cancer Cells Through a Synergistic Action of Luteolin and Paclitaxel.

BACKGROUND AND AIM:  According to reports on cancer incidence in 2020, breast cancer became the leading malignancy among women worldwide. This multistep disease involves genetic and environmental factors. Paclitaxel, a naturally occurring antimitotic substance, is a widely used chemotherapeutic drug for treating various human malignancies, including breast cancer. However, its major drawback is its extensive toxicity. This limitation can be mitigated through combination therapy with natural products like luteolin. Studies suggest that luteolin has anticancer properties, as it inhibits cancer cell growth and induces apoptosis in breast, lung, and colon cancers. This study aims to investigate the synergistic anticancer effects of combining luteolin and paclitaxel on breast cancer cells. METHODS: Breast cancer cell line (MDA-MB-231) was utilized for this study. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was then conducted to check the cell viability. This was followed by a morphology study conducted under a phase contrast microscope. Morphological analysis revealed pronounced cell shrinkage and membrane blebbing, indicative of apoptosis when treated with the combination at their IC50 values. Gene expression results further confirmed the anticancer properties by showing significant downregulation of the B-cell lymphoma-2 (BCL-2) anti-apoptotic gene. These findings suggest that the luteolin-paclitaxel combination exerts a synergistic effect, enhancing anticancer activity in breast cancer cells. Reverse transcriptase polymerase chain reaction (RT-PCR) was done to analyze the genes involved in apoptosis. Finally, the data collected was statistically analyzed to confirm the reliability of the study. RESULTS: The combination of 1 µM/ml of paclitaxel and increasing concentrations of luteolin showed a great percentage of reduction in cell viability and the IC50 value of luteolin concentration was around 40 µM/ml. The morphology study revealed that the cancer cells showed shrinkage and blebbing on treatment with 40 µM/ml. At the same IC50 concentration, the combination of luteolin and paclitaxel resulted in a significant downregulation of BCL-2 mRNA expression in breast cancer cells compared to luteolin alone. CONCLUSION: The combination of paclitaxel and luteolin has a synergistic effect on breast cancer cells and shows potential as a treatment for various cancers. Given these promising results, the paclitaxel and luteolin combination could be developed into a potent therapeutic strategy for treating various cancers. Future research should include in vivo studies to further assess the therapeutic potential and safety profile of this combination.