Molecular mechanism of drug induced apoptosis and chemoresistance in estrogen receptor alpha +/- breast cancer cell lines: mcf-7 and mda-mb-231

It was recently shown that inhibition or downregulation of Bcl-2 represents a new therapeutic approach to by-pass chemoresistance mechanism in cancer cells. Therefore, we explored the potential of this approach in breast cancer cells; MCF-7 (drug-sensitive; p53 wild type) and MDA-MB-231 (drug-insensitive; p53 mutant). Cisplatin and paclitaxel induced apoptosis in a dose-dependent manner in both cell lines. Furthermore, silencing of Bcl-2 remarkably increased cisplatin and paclitaxel induced apoptosis. Dose dependent induction of apoptosis by cisplatin and paclitaxel was enhanced by the pre-treatment of these cells with HA14-1, a Bcl-2 inhibitor. Although the effect of cisplatin on cell death was significant in MCF-7 and MDA-MB- 231, paclitaxel was less potent only in MDA-MB-231 cells. To further understand the distinct role of drugs in breast cancer cells which were pre-treated with HA14-1, changes in mitochondrial membrane potential, caspase activation, and Bcl-2 family protein levels, generation of reactive oxygen species and lipid peroxidation were studied. The apoptotic effect of cisplatin with or without HA14- 1 pre-treatment was shown to be caspase-dependent in both cell lines. While proapoptotic Bcl-2 proteins (Bax, Puma, Bad) were found to be up-regulated, Bcl-2 and Bcl-xL were down-regulated when cells were pre-treated with HA14-1 followed by cisplatin or paclitaxel. MCF-7 and MDA-MB-231 cells overexpressing Bcl-2 displayed different responses upon drug-treatment. Although cisplatin could still induce apoptosis in Bcl-2 overexpressing MCF-7 cells by promoting pro-apoptotic Bcl-2 family members, Bcl-2 overexpression abrogated paclitaxel induced apoptosis in MCF-7 and MDA-MB-231 breast cancer cells, respectively. In conclusion, our findings suggest two important implications for understanding cisplatin and paclitaxel induced apoptosis mechanism and the potential role of Bcl-2 in this apoptotic pathway. First, the potentiating effect of Bcl-2 inhibitor (HA14-1) is drug and cell type specific and may not only depend on the inhibition of Bcl-2. Importantly, alteration of other pro-apoptotic or anti-apoptotic Bcl-2 family members may dictate the apoptotic response when HA14-1 is combined with chemotherapeutic drugs. Second, cisplatin activated a p53- regulated pro-apoptotic pathway to overcome Bcl-2 mediated resistance. These insights may be useful for the development of novel treatments for cancer cells overexpressing anti-apoptotic Bcl-2 proteins

Applications of Aptamers in Cancer Therapy

Aptamers are small and specific oligonucleotides [RNA or single-strand DNA (ssDNA)] with a high binding affinity against target protein. In vitro selection process of aptamer by selective evolution of ligands by exponential enrichment (SELEX) has been invented in 1990 by Larry Gold and Jack Szostak. SELEX is a random amplification of target protein with combined oligonucleotide libraries and selection of synthesized aptamer by magnetic beads, affinity chromatography, and capillary electrophoresis-based methods. According to their low molecular weight, non-immunogenic feature in vivo, low production cost, high thermal stability, increase in production potential, and ample of modification capacities, aptamers are becoming essential medical tools for diagnosis and treatment of diseases such as macular degeneration, hemophilia, heart disease, and various cancer types. The therapeutic potential of aptamers, with high binding affinity against carcinogenesis-associated growth factors, receptors, or proteins frequently overexpressed in specific cancers such as prostate, breast, colon, lung, leukemia, hepatocellular, and cervical carcinoma. The strategies for aptamer-based drugs in cancer therapy design/modify aptamers against cancer biomarkers, accelerate immunotherapy targeting immune system, and increase the drug delivery in cancer cells. In conclusion, aptamers are promising candidate drugs due to their antiproliferative effect on cancer cells and the drug delivery systems during cancer chemotherapy

Aging-Related Diseases and Autophagy

Autophagy is fundamental, evolutionary conserved physiological process at molecular level which targets long-lived cytosolic proteins and organelles to be recycled through lysosomal degradation. Diminished autophagic activity caused cellular stress in many organisms following aging, and inhibition of autophagy in model organisms causes degenerative changes and pathologic diseases observed with high incidence ratio generally in older ages. Consequently the delayed senescence or increased longevity in model organisms often stimulate autophagy, and autophagy inhibition compromises anti-aging effects. The cytoprotective function of autophagy is presented in various human diseases such as lung, liver, cardiovascular diseases, neurodegeneration, myopathies, cancer, stroke, infections and metabolic diseases which are found associated with autophagic targets. These pathologies are defined with their age-dependent characteristics, is not fully understood that how autophagy network regulates metabolism and may cause diseases in age-related manner. In this book chapter, we are going to discuss the autophagy and aging relationship in three different parts. In the first section autophagy and aging relationship is going to be presented through explaining responsible signalling network. The autophagy and age-related neurological disorders, genetic basis of age-dependent diseases and the functional role of autophagy is going to be discussed in the second and third part of the chapter

Breast Cancer and Flavonoids as Treatment Strategy

Breast cancer is the most prevalent cancer type among women. Despite recent progress in early detection and therapeutic strategies, the rate of mortality is increasing. Anti-estrogens or aromatase inhibitors are preferred to treat the women diagnosed with estrogen-receptor (ER) positive tumors. However, breast tumors usually show intra-tumoral heterogeneity with ER-positive and -negative cells. The advanced breast cancer cells lose the estrogen responsiveness and become aggressive by developing new strategies for rapid proliferation such as mutations in cell cycle machinery. New promising drugs are still being investigating against these types of tumors especially to overcome acquired resistance against chemotherapeutic drugs; however, a successful treatment for metastatic tumors is still unclear. Flavonoids, with various pharmacological activities, are plant or fungus secondary metabolites present in human diet. In plants, beside their role in pigmentation, they may also act as messengers, regulators and cell cycle inhibitors. Therefore, they are being tested in ovarian, cervical as well as breast cancer. Due to the positive correlation between flavonoids-rich diet and lower risk of cancer, flavonoids are referred as chemopreventive agents. The current chapter emphasizes the therapeutic potential of flavonoids and their synthetic analogues as anti-cancer agents in breast cancer providing new insights into the molecular mechanisms

Specific c-Jun N-Terminal Kinase Inhibitor, JNK-IN-8 Suppresses Mesenchymal Profile of PTX-Resistant MCF-7 Cells through Modulating PI3K/Akt, MAPK and Wnt Signaling Pathways

Paclitaxel (PTX) is a widely used chemotherapeutic agent in the treatment of breast cancer, and resistance to PTX is a common failure of breast cancer therapy. Therefore, understanding the effective molecular targets in PTX-resistance gains importance in identifying novel strategies in successful breast cancer therapy approaches. The aim of the study was to investigate the functional role of PTX resistance on MCF-7 cell survival and proliferation related to PI3K/Akt and MAPK pathways. The generated PTX-resistant (PTX-res) MCF-7 cells showed enhanced cell survival, proliferation, and colony formation potential with decreased cell death compared to wt MCF-7 cells. PTX-res MCF-7 cells exhibited increased motility profile with EMT, PI3K/Akt, and MAPK pathway induction. According to the significant SAPK/JNK activation in PTX-res MCF-7 cells, specific c-Jun N-terminal kinase inhibitor, JNK-IN-8 is shown to suppress the migration potential of cells. Treatment of JNK inhibitor suppressed the p38 and SAPK/JNK and Vimentin expression. However, the JNK inhibitor further downregulated Wnt signaling members in PTX-res MCF-7 cells. Therefore, the JNK inhibitor JNK-IN-8 might be used as a potential therapy model to reverse PTX-resistance related to Wnt signaling

microRNA 1307 Is a Potential Target for SARS-CoV-2 Infection: An <i>in Vitro</i> Model

microRNAs (miRs) are proposed as critical molecular targets in SARS-CoV-2 infection. Our recent in silico studies identified seven SARS-CoV-2 specific miR-like sequences, which are highly conserved with humans, including miR-1307-3p, with critical roles in COVID-19. In this current study, Vero cells were infected with SARS-CoV-2, and miR expression profiles were thereafter confirmed by qRT-PCR. miR-1307-3p was the most highly expressed miR in the infected cells; we, therefore, transiently inhibited its expression in both infected and uninfected cells. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) cell proliferation assay assessed cell viability following SARS-CoV-2 infection, identifying that miR-1307 expression is inversely correlated with cell viability. Lastly, changes in miR-1307-dependent pathways were analyzed through a detailed miRNOME and associated in silico analysis. In addition to our previously identified miRs, including miR-1307-3p, the upregulation of miR-193a-5p, miR-5100, and miR-23a-5p and downregulation of miR-130b-5p, miR34a-5p, miR-505-3p, miR181a-2-3p, miR-1271-5p, miR-598-3p, miR-34c-3p, and miR-129-5p were also established in Vero cells related to general lung disease-related genes following SARS-CoV-2 infection. Targeted anti-miR-1307-3p treatment rescued cell viability in infection when compared to SARS CoV-2 mediated cell cytotoxicity only. We furthermore identified by in silico analysis that miR-1307-3p is conserved in all SARS-CoV-2 sequences/strains, except in the BA.2 variant, possibly contributing to the lower disease severity of this variant, which warrants further investigation. Small RNA seq analysis was next used to evaluate alterations in the miRNOME, following miR-1307-3p manipulation, identifying critical pathobiological pathways linked to SARS-CoV-2 infection-mediated upregulation of this miR. On the basis of our findings, miRNAs like miR-1307-3p play a critical role in SARS-CoV-2 infection, including via effects on disease progression and severity

Small inhibitor of Bcl-2, HA14-1, selectively enhanced the apoptotic effect of cisplatin by modulating Bcl-2 family members in MDA-MB-231 breast cancer cells

Inhibition or downregulation of Bcl-2 represents a new therapeutic approach to by-pass chemoresistance in cancer cells. Therefore, we explored the potential of this approach in breast cancer cells. Cisplatin and paclitaxel induced apoptosis in a dose-dependent manner in MCF-7 (drug-sensitive) and MDA-MB-231 (drug-insensitive) cells. Furthermore, when we transiently silenced Bcl-2, both cisplatin and paclitaxel induced apoptosis more than parental cells. Dose dependent induction of apoptosis by drugs was enhanced by the pre-treatment of these cells with HA14-1, a Bcl-2 inhibitor. Although the effect of cisplatin was significant on both cell lines, the effect of paclitaxel was much less potent only in MDA-MB-231 cells. To further understand the distinct role of drugs in MDA-MB-231 cells pretreated with HA14-1, caspases and Bcl-2 family proteins were studied. The apoptotic effect of cisplatin with or without HA14-1 pre-treatment is shown to be caspase-dependent. Among pro-apoptotic Bcl-2 proteins, Bax and Puma were found to be up-regulated whereas Bcl-2 and Bcl-x(L) were down-regulated when cells were pretreated with HA14-1 followed by paclitaxel or cisplatin. Enforced Bcl-2 expression in MDA-MB-231 cells abrogated the sensitizing effect of HA14-1 in cisplatin induced apoptosis. These results suggest that the potentiating effect of HA14-1 is drug and cell type specific and may not only depend on the inhibition of Bcl-2. Importantly, alteration of other pro-apoptotic or anti-apoptotic Bcl-2 family members may dictate the apoptotic response when HA14-1 is combined with chemotherapeutic drugs

Extracts of Cistus creticus cultivated at different salinity levels exhibit promising therapeutic potential for pancreatic cancer cell lines

Background: Cistus creticus L. subsp. creticus is a characteristic species commonly distributed in the Mediterranean region. It has been proved to contain antibacterial, antifungal, anti-inflammatory, and anti-cancer traits. Purpose: In this study, C. creticus plants were grown in hydroponic culture under different salinity levels (0, 30, or 60 mM NaCl) and were extracted using two other solvents (70 % ethanol and distilled water). The effect of salinity on plant growth and mineral homeostasis, biochemical content of extracts and therapeutic impact on cell death and cell survival in pancreatic cancer cell lines were studied. Methods: Pancreatic cancer cell lines (PANC-1, MIA PaCa-2, and AsPC-1) were treated with different C. creticus extracts were grown, and cell survival, autophagy and apoptosis signaling pathways were measured. Results: Especially, ethanol extraction of C. creticus showed a remarkably therapeutic effect on cell death and cell survival. Ethanol extracts have higher polyphenolic content and are more effective than water extracts. It has been found that C. creticus is an important mitochondria-targeted antioxidant in pancreatic cancer cells. Conclusion: This research could be important to evaluate how stress physiology relates to the chemical composition and the therapeutic effectiveness of C. creticus