Shaymaa M. M. Yahya

Background This study sought to uncover novel targets for fighting drug resistance in breast cancer therapy, such as miRNA that target ABC transporters and decrease the oncogenic activity of breast cancer cells. Methods MiRNA ABC transporter targets were predicted using in silico methods after evaluating the ABC transporter route. In addition, miRNA expression, gene annotation, and gene ontology concepts were investigated. The expression levels of ABC transporter genes and miRNAs were determined using QRT-PCR. Dual luciferase activity was assessed to establish the precise ABC transporter-specific interaction. Flowcytometric analysis was used to detect doxorubicin uptake in miR-4330-treated cells, as well as cell cycle analyses. The spreading capacity of miR-4330-upregulated cells was investigated. Results The current study identified miR-4330 as an anti-drug and anti-oncogenic molecule in both estrogen receptor positive and negative breast cancer cells. Furthermore, the study identified the mechanism by which miR-4330 restoration reduces drug resistance by drastically decreasing ABCG2 expression (fold change of -0.25), which was accompanied by a considerable increase in doxorubicin accumulation or influx in both cell lines. Furthermore, miRNA-4330 restoration resulted in a considerable reduction in metastatic and spreading capabilities in both cell types. Conclusion The current findings identified miR-4330 restoration as a tool for overcoming acquired drug resistance and reducing cancer activity, which might be used to improve therapeutic strategies and treatment regimens for breast cancer patients. miR-4330 may be useful as a diagnostic and prognostic marker for drug-resistant breast cancer.

Purpose: Breast cancer is one of the leading types of cancer diagnosed in women. Despite the improvements in chemotherapeutic cure strategies, drug resistance is still an obstacle leading to disease aggressiveness. The small non-coding RNA molecules, miRNAs, have been implicated recently to be involved as regulators of gene expression through the silencing of mRNA targets that contributed to several cellular processes related to cancer metastasis. Hence, the present study aimed to investigate the beneficial role and mechanism of miRNA-34a-based gene therapy as a novel approach for conquering drug resistance mediated by ATP-binding cassette (ABC) transporters in breast cancer cells, besides exploring the associated invasive behaviors. Material and Methods: Bioinformatics tools were used to predict miRNA ABC transporter targets by tracking the ABC transporter pathway. After the establishment of drug-resistant breast cancer MCF-7 and MDA-MB-231 sublines, cells were transfected with the mimic or inhibitor of miRNA-34a-5p. The quantitative expression of genes involved in drug resistance was performed by QRT-PCR, and the exact ABC transporter target specification interaction was confirmed by dual-luciferase reporter assay. Furthermore, flow cytometric analysis was utilized to determine the ability of miRNA-34a-treated cells against doxorubicin uptake and accumulation in cell cycle phases. The spreading capability was examined by colony formation, migration, and wound healing assays. The apoptotic activity was estimated as well. Results: Our findings firstly discovered the mechanism of miRNA-34a-5p restoration as an anti-drug-resistant molecule that highly significantly attenuates the expression of ABCC1 via the direct targeting of its 3′- untranslated regions in resistant breast cancer cell lines, with a significant increase of doxorubicin influx by MDA-MB-231/Dox-resistant cells. Additionally, the current data validated a significant reduction of metastatic potentials upon miRNA-34a-5p upregulation in both types of breast cancer-resistant cells. Conclusion: The ectopic expression of miRNA-34a ameliorates the acquired drug resistance and the migration properties that may eventually lead to improved clinical strategies and outcomes for breast cancer patients. Additionally, miRNA-34a could be monitored as a diagnostic/prognostic biomarker for resistant conditions.

Hepatocellular carcinoma (HCC) is the major lethal primary liver malignant worldwide. Although, melatonin has various antitumor bioactivities; there is a requirement for more investigations to elucidate the not discussed effects, and the controversial responses of the treatment with melatonin on targets mediated in HCC. To achieve the aim of the present study, HCC-HepG2 cells were treated with different concentrations of melatonin at various time intervals. The selected minimal proliferation inhibition doses of melatonin were then incubated with cells to examine the arresting effect of melatonin on dividing cells using flow cytometry. Furthermore, the molecular patterns of genes that contributed to apoptosis, drug resistance development, antioxidation, and melatonin crossing were quantified by qRT-PCR. Additionally, the Human inflammation antibody array membrane (40 targets) was used to check the anti-inflammatory effect of melatonin. Our results validated that, melatonin shows anti-proliferative action through preserving cells in G0/G1 phase (P < 0.001) that is associated with a highly significant increase in the expression level of the P53 gene (P < 0.01). On contrary, as a novelty, our data recorded decreases in expression levels of genes involved in the pro-apoptotic pathway; with a significant increase (P < 0.05) in the expression level of an anti-apoptotic gene, Bcl2. Interestingly, we detected observed increases in the expression levels of genes responsible for conferring drug resistance including ABCB1, ABCC1, and ABCC5. Our study proved the anti-inflammatory activity of 1 mM melatonin in HCC-HepG2 cells. Accordingly, we can conclude that melatonin facilitates the anti-proliferation of cells at doses of 1 mM, and 2.5 mM after 24 h. This action is initiated through cell cycle arrest at G0/G1 phase via increasing the expression of P53, but independently on apoptosis. Collectively, melatonin is an effective anti-inflammatory and anti-proliferative promising therapy for the treatment of HCC. However, its consumption should be cautious to avoid the development of drug resistance and provide a better treatment strategy.

Hepatocellular carcinoma (HCC) is the major life-threatening primary liver malignancy in both sexes all over the world. Unfortunately, the majority of patients are diagnosed at later stages because HCC does not elicit obvious symptoms during its early incidence. Consequently, most individuals escape the first-line HCC treatments and are treated with chemotherapy. Regrettably, the therapeutic outcomes for those patients are usually poor because of the development of multidrug resistance phenomena. Furthermore, most anti-HCC therapies cause severe undesired side effects that notably interfere with the life quality of such patients. Accordingly, there is an important need to search for an alternative therapeutic drug or adjuvant which is more efficient with safe or even minimal side effects for HCC treatment. Melatonin was recently reported to exert intrinsic antitumor activity in different cancers. However, the regulatory pathways underlying the antitumor activity of melatonin are poorly understood in resistant liver cells. Furthermore, a limited number of studies have addressed the therapeutic role of melatonin in HCC cells resistant to doxorubicin chemotherapy. In this study, we investigated the antitumor effects of melatonin in doxorubicin-resistant HepG2 cells and explored the regulatory pivotal targets underlying these effects. To achieve our aim, an MTT assay was used to calculate the 50% inhibitory concentration of melatonin and evaluate its antiproliferative effect on resistant cells. Additionally, qRT-PCR was used to quantify genes having a role in drug resistance phenotype (ABCB1, ABCC1, ABCC2, ABCC3, ABCC4, ABCC5, and ABCG2); apoptosis (caspases-3, and -7, Bcl2, Bax, and p53); anti-oxidation (NRF2); expression of melatonin receptors (MT1, MT2, and MT3); besides, programmed death receptor PD-1 gene. The active form of the caspase-3 enzyme was estimated by ELISA. A human inflammatory antibody membrane array was employed to quantify forty inflammatory factors expressed in treated cells. We observed that melatonin inhibited the proliferation of doxorubicin-resistant HepG2 cells in a dose-dependent manner after 24-h incubation time with a calculated IC₅₀ greater than 10 mM (13.4 mM), the expression levels of genes involved in drug resistance response (ABCB1, ABCC1, ABCC5, and ABCG2) were downregulated. Also, the expression of caspase-3, Caspase-7, NRF2, and p53 genes were expressed at higher levels as compared to control (DMSO-treated cells). An active form of caspase-3 was confirmed by ELISA. Moreover, the anti-inflammatory effect of melatonin was detected through the calculated fold change to control which was reduced for various mediators that have a role in the inflammation pathway. The current findings introduce melatonin as a promising anti-cancer treatment for human-resistant HCC which could be used in combination with current chemotherapeutic regimens to improve the outcome and reduce the developed multidrug resistance.

Background: Colorectal cancer is among the leading causes of death worldwide. The incidence of deaths is expected to be 11.4 million in 2030.Objective: We aimed to evaluate the in vitro and in vivo antioxidant and antitumor activities of a novel Bithiophene- Fluorobenzamidine (BFB) against DMH-induced colorectal cancer in rats.Methods: The antiproliferative activity of BFB against HCT-116 colon cancer cells and apoptotic genes was assessed. In vivo study was also conducted in which 80 adult male rats were divided into 5 groups; control, BFB, and the other 3 groups were injected with DMH (20mg/kg, s.c., for 9 weeks). Group 4 was injected with 5 doses of cisplatin (2.5mg/kg, i.p over 21 weeks) and group 5 was injected with 3 doses/week of BFB (2.5mg/kg, i.p, for 21 weeks).Results: BFB exhibited weak to moderate in vitro antioxidant activity. It had a strong antiproliferative activity with IC50 ~0.3µg/ml. BFB induced extrinsic apoptosis through the upregulation of FasL, TRAL, p53 and caspase-8, and intrinsic apoptosis through the downregulation of Bcl-2 and survivin. BFB decreased the tumor incidence, multiplicity and size and improved the decreased body weight. BFB also ameliorated the functions of kidney and liver and antioxidants deteriorated by DMH. BFB significantly improved the pathological changes caused by DMH in colon tissues.Conclusion: BFB showed a very promising antitumor activity against colorectal cancer induced by DMH in rats without causing hepato- or nephrotoxicity.

Background: Chemotherapeutic drugs have high toxicity associated with undesirable side-effects. Now, natural products are the most important anti-cancer agents because of their low toxicity and potential effectiveness. Methods: The half maximal inhibitory concentration (IC50) of amygdalin, naringenin and ellagic acid against breast, colon, and liver cell lines was estimated. The antimutagenic, free radical-, superoxide radical-, and hydroxyl radical- scavenging activities of these phytochemicals were measured. The expression of p53, bid, bax, bcl2, and caspases 9, 3, and 7 was measured by quantitative real-time polymerase chain reaction (qRT-PCR) in breast and liver cells. In addition, the active Caspase 3 protein was estimated in liver cells. Results: Ellagic acid showed the highest antioxidant and antiproliferative activities. Amygdalin and naringenin with low and moderate antioxidant profiles showed a corresponding low and moderate cytotoxicity against cancer cell lines, respectively. Naringenin and ellagic acid had a significant antimutagenic activity which was detected by the Salmonella test. Ellagic acid offered a much better antimutagenic activity than naringenin. The apoptotic pathway evoked by ellagic acid in HepG2 and MCF-7 cells was investigated. The results showed that a caspase-dependent and a caspase-independent apoptosis occurred in MCF-7 and HepG2, respectively. Conclusion: The antimutagenic/antioxidant properties are well correlated with the antiproliferative activity of the phytochemicals investigated. This study proved that some easy, quick and cheap assays could predict the antiproliferative activity of many nutraceuticals. Finally, this platform could help in the discovery of new anticancer agents where hundreds of compounds are investigated in the pipeline of drug discovery.

Hepatocellular carcinoma (HCC) was accompanied by high incidence of morbidity and mortality worldwide. Apoptosis is a vital biological process playing a critical role in cancer. Besides, toll like receptors were reported to regulator the innate immune response against cancer development. Exopolysaccharides (EPSs) derived from marine bacteria were reported to have a potential biological importance. This work aimed to elucidate the antitumor effects of newly isolated EPSs against HepG2 cells. Moreover, their effects on some apoptotic markers and TLRs were followed. Isolated EPSs were tested for their cytotoxic effects in a previous study and the most promising; MSA1, E4, MGA2, SGA3, and NRC7 EPSs were subjected to molecular analysis to investigate their pro-apoptotic effects, in addition to their effects on TLR2 and TLR-9 using quantitative real time RT-PCR. And the most cytotoxic and pro-apoptotic EPS; MSA1 were subjected to antibody array analysis to investigate a panel of 43 apoptotic proteins. All isolated EPSs produced a positive role in regulating the apoptotic gene and increasing the TLRs expression in different manners. However, the most promising EPS was MSA1. It showed pro-apoptotic effects on gene and protein levels, besides its up-regulation of TLRs.

Hepatocellular carcinoma (HCC) is one of the foremost causes of cancer related morbidity worldwide. An increasing number of studies have confirmed that microRNAs play an important role in the development, progression and metastasis of HCC. From those important miRNAs are miR-98 and miR-214. This study were conducted to explore the effect of these two miRNAs on some apoptotic and angiogenic genes namely, BCL-2, survivin, CCND1, CDC2, P53 and P21, VEGF, Hif-1α, MMP-2, MMP-9, Ang-1, Ang-2, and FGF-1. miRNAs mimics and inhibitors transfection was used to investigate the role of both studied molecules in apoptosis and angiogenesis in HepG2 cells. QRT-PCR was used for Quantitative gene and miRNA expression analyses. The study revealed that miR-98 could serve as a pro-apoptotic factor through the upregulation of P53 gene expression levels. Besides, the anti-angiogenic effect of this miRNA was evident through the down regulation of Ang-1 and FGF-1 genes. Meanwhile, miR-214 showed a pro-apoptotic role and anti-angiogenic effects. These effects were verified through the significant down regulation of BCL-2, CDC2, VEGF, Ang-1 and MMP-2. These results introduced a possible positive role played by both miR-98 and miR-214 on some pro-apoptotic and anti-angiogenic genes.