Sherif Abdelhamed

Chrysin overcomes TRAIL resistance of cancer cells through Mcl-1 downregulation by inhibiting STAT3 phosphorylation

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) selectively kills various types of cancer cells without harming normal cells, but TRAIL resistance has been frequently observed in cancer cells. Propolis (bee glue) is a material collected from various plants by honeybees and is a rich source of bioactive compounds, including the natural flavonoid chrysin, which possesses multiple anticancer effects. We investigated the mechanism underlying the TRAIL sensitization effect of chrysin, which is a major constituent of Thai propolis, in human lung and cervical cancer cell lines. Propolis extract and chrysin sensitizes A549 and HeLa human cancer cell lines to TRAIL-induced apoptosis. The TRAIL sensitization effect of chrysin is not mediated by inhibition of TRAIL-induced NF-κB activation or by glutathione depletion. Immunoblot analysis using a panel of anti-apoptotic proteins revealed that chrysin selectively decreases the levels of Mcl-1 protein, by downregulating Mcl-1 gene expression as determined by qRT-PCR. The contribution of Mcl-1 in TRAIL resistance was confirmed by si-Mcl-1 knockdown. Among signaling pathways that regulate Mcl-1 gene expression, only constitutive STAT3 phosphorylation was suppressed by chrysin. The proposed action of chrysin in TRAIL sensitization by inhibiting STAT3 and downregulating Mcl-1 was supported by using a STAT3‑specific inhibitor, cucurbitacin-I, which decreased Mcl-1 levels and enhanced TRAIL-induced cell death, similar to that observed with chrysin treatment. In conclusion, we show the potential of chrysin in overcoming TRAIL resistance of cancer cells and elucidate its mechanism of action.

Berberine enhances tumor necrosis factor-related apoptosis-inducing ligand-mediated apoptosis in breast cancer.

Berberine (BBR) has been used for the treatment of bacterial and fungal infections and also for cancer-associated symptoms such as diarrhea. Furthermore, it has been reported that BBR may have direct antitumor effects. Although evidence supports the theory that tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is a promising candidate for treating cancer, its usage may be limited due to the resistance to the TRAIL-induced apoptosis of cancer cells. In the present study, the effect of BBR on TRAIL-induced antitumor effects was investigated in vitro using recombinant TRAIL and in vivo using a 4T1 murine breast cancer model in combination with anti-DR5 (death-inducing TRAIL receptor) monoclonal antibody therapy. BBR sensitized human breast cancer cell lines to TRAIL-mediated apoptosis in vitro. The combination of BBR and recombinant TRAIL significantly activated caspase-3 and PARP cleavage in TRAIL-resistant MDA-MB-468 cells. Furthermore, BBR in combination with TRAIL more effectively induced apoptosis compared with coptisine (COP), which is structurally related to BBR. In a murine 4T1 breast cancer model, BBR treatment enhanced the efficacy of anti-DR5 antibody therapy against primary tumor growth and lung metastasis. Thus, BBR may become a new adjuvant for overcoming the resistance of cancer cells to TRAIL/DR5-mediated therapy.

Antiausterity Agents from Uvaria dac and Their Preferential Cytotoxic Activity against Human Pancreatic Cancer Cell Lines in a Nutrient-Deprived Condition

Human pancreatic cancer cell lines are known for their inherent tolerance to nutrition starvation, which enables them to survive under a hypovascular (austerity) tumor microenvironment. The search for agents that preferentially retard the survival of cancer cells under low nutrition conditions (antiausterity agent) is a novel approach to anticancer drug discovery. In this study, it was found that a dichloromethane extract of the stem of Uvaria dac preferentially inhibited PANC-1 human pancreatic cancer cells survival under nutrition-deprived conditions at a concentration of 10 μg/mL. Workup of this bioactive extract led to the discovery of (+)-grandifloracin (8) as a potent antiausterity agent as evaluated in a panel of four human pancreatic cancer cell lines, PANC-1 (PC(50), 14.5 μM), PSN-1 (PC(50), 32.6 μM), MIA PaCa-2 (PC(50), 17.5 μM), and KLM-1 (32.7 μM). (+)-Grandifloracin (8) has been isolated from a natural source for the first time. Its absolute stereochemistry was established by single-crystal X-ray crystallography and circular dichroism spectroscopic analysis. In addition to this, seven other new highly oxygenated cyclohexene derivatives, named uvaridacanes A (1) and B (2), uvaridacols A-D (3, 4, 6, 7), and uvaridapoxide A (5), were also isolated and structurally characterized.

AKT-STAT3 Pathway as a Downstream Target of EGFR Signaling to Regulate PD-L1 Expression on NSCLC cells

While cancer development and progression can be controlled by cytotoxic T cells, it is also known that tumor-specific CD8+T cells become functionally impaired by acquiring a group of inhibitory receptors known as immune checkpoints. Amongst those, programmed death-1 (PD-1) is one of the most recognized negative regulators of T cell function. In non-small lung cancers (NSCLCs), the aberrant activation of epidermal growth factor receptor (EGFR) is known to induce PD-L1 expression and further the treatment with gefitinib, a tyrosine kinase inhibitor (TKI) for EGFR, decrease the expression of PD-L1 on NSCLC. Given the acquired resistance to gefitinib treatment frequently observed by developing secondary-site mutations limiting its efficacy, it is important to understand the downstream mechanism of activated-EGFR signaling for regulating PD-L1 in NSCLC. In this study, we demonstrated that AKT-STAT3 pathway could be a potential target for regulating the surface expression of PD-L1 on NSCLCs with aberrant EGFR activity and, further, the inhibition of AKT or STAT3 activity could down-regulate the expression of PD-L1 even in gefitinib-resistant NSCLCs. These results highlight an importance of AKT-STAT3 pathway as a promising target for potentiating anti-tumor immune responses by regulating PD-L1 expression on cancer cells with aberrant EGFR activity.

Identification of plant extracts sensitizing breast cancer cells to TRAIL

Triple-negative breast cancer (TNBC) is an aggressive heterogeneous cancer subgroup with a higher rate of distant recurrence and a poorer prognosis compared to other subgroups. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is an attractive molecule that induces cell death in various tumor cells without causing cytotoxicity to normal cells; however, primary or acquired resistance to TRAIL often limits its efficacy in cancer patients. To develop combination therapies to improve TRAIL efficacy and/or to overcome the resistant mechanism, we screened 138 medicinal plant extracts against TRAIL-sensitive and -insensitive TNBC cell lines, MDA-MB-231 and MDA-MB-468. Among them, 5 plant extracts, Uvaria dac, Artemisia vulgaris, Cortia depressa, Dichasia bengalensis and Cinnamomum obtusifolium did not cause apparent cytotoxicity (<20%) as a single regimen, but showed significant synergistic effects in combination with TRAIL against both cell lines. Moreover, Uvaria dac, Artemisia vulgaris and Cinnamomum obtusifolium were found to suppress the phosphorylation of p65 that is involved in TRAIL-resistant mechanisms. These observations suggest that the identified plant extracts in combination with TRAIL could lead to potential therapeutic benefits for cancer patients in the clinical setting.

Uvaridacols E−H, Highly Oxygenated Antiausterity Agents from Uvaria dac

Chemical investigation of the stems of Uvaria dac yielded four new highly oxygenated cyclohexene derivatives named uvaridacols E-H (1-4). Their structures were established through NMR and circular dichroism spectroscopic analysis. Uvaridacols E (1), F (2), and H (4) displayed weak preferential cytotoxicity against PANC-1 human pancreatic cancer cells under nutrition-deprived conditions in a concentration-dependent manner, without causing toxicity in normal nutrient-rich conditions.

Role of tyrosine kinase-independent phosphorylation of EGFR with activating mutation in cisplatin-treated lung cancer cells

Epidermal growth factor receptor (EGFR) mutation is one of the hallmarks of cancer progression and resistance to anticancer therapies, particularly non-small cell lung carcinomas (NSCLCs). In contrast to the canonical EGFR activation in which tyrosine residues are engaged, we have demonstrated that the non-canonical pathway is triggered by phosphorylation of serine and threonine residues through p38 and ERK MAPKs, respectively. The purpose of this study is to investigate the role of non-canonical EGFR pathway in resistance mechanism against cisplatin treatment. Wild type and mutated (exon 19 deletion) EGFR-expressing cells responded similarly to cisplatin by showing MAPK-mediated EGFR phosphorylation. It is interesting that internalization mechanism of EGFR was switched from tyrosine kinase-dependent to p38-dependent fashions, which is involved in a survival pathway that counteracts cisplatin treatment. We therefore introduce a potential combinatorial therapy composed of p38 inhibition and cisplatin to block the activation of EGFR, therefore inducing cancer cell death and apoptosis.

Bufadienolides Overcome TRAIL Resistance in Breast Cancer Cells via JAK-STAT Pathway

Aim and objective: Although the clinical significance of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has been elucidated, primary or acquired resistance to TRAIL limits its efficacy in cancer patients. This study aimed to investigate bufadienolides as promising candidates to combine with TRAIL and to determine the pathway involved in overcoming TRAIL resistance in breast cancer cells. Methods: The effects of bufadienolides, cinobufagin and cinobufotalin, in combination with TRAIL were examined in TRAIL-sensitive and TRAIL-resistant breast cancer, MDA-MB-231 and MDA-MB-468 cells, respectively. Cell viability was determined using count and viability assays. Apoptotic cell death was detected by annexin-V/7-AAD staining. Silencing of MCL-1 expression was established using MCL-1 siRNA and protein expression was assessed by Western blot analysis. STAT3 localization was determined using immunofluorescence staining. Results: The combination of bufadienolides with TRAIL significantly reduced cell viability in both TRAIL-sensitive and TRAIL-resistant breast cancer cells in vitro. Bufadienolides overcame TRAIL resistance by downregulating MCL- 1, an anti-apoptotic protein, through the JAK-STAT pathway. The combination of TRAIL with knock down of MCL-1 or JAK-STAT inhibitor, JSI-124, could synergistically induce apoptosis similar to bufadienolides. Conclusion: Bufadienolides enhance TRAIL-induced apoptosis in breast cancer cells through Mcl-1 downregulation via JAK-STAT pathway. Targeting MCL-1 or JAK2 could be one of the strategies to overcome breast cancer cell resistance to TRAIL. Bufadienolides, through JAK2/STAT3/MCL-1 inhibition, could be considered as promising cancer therapy agents in combination with TRAIL.

Retrospective screening of microarray data to identify candidate IFN-inducible genes in a HTLV-1 transformed model

HuT-102 cells are considered one of the most representable human T-lymphotropic virus 1 (HTLV-1)-infected cell lines for studying adult T-cell lymphoma (ATL). In our previous studies, genome-wide screening was performed using the GeneChip system with Human Genome Array U133 Plus 2.0 for transforming growth factor-β-activated kinase 1 (TAK1)-, interferon regulatory factor 3 (IRF3)- and IRF4-regulated genes to demonstrate the effects of interferon-inducible genes in HuT-102 cells. Our previous findings demonstrated that TAK1 induced interferon inducible genes via an IRF3-dependent pathway and that IRF4 has a counteracting effect. As our previous data was performed by manual selection of common interferon-related genes mentioned in the literature, there has been some obscure genes that have not been considered. In an attempt to maximize the outcome of those microarrays, the present study reanalyzed the data collected in previous studies through a set of computational rules implemented using ‘R’ software, to identify important candidate genes that have been missed in the previous two studies. The final list obtained consisted of ten genes that are highly recommend as potential candidate for therapies targeting the HTLV-1 infected cancer cells. Those genes are ATM, CFTR, MUC4, PARP14, QK1, UBR2, CLEC7A (Dectin-1), L3MBTL, SEC24D and TMEM140. Notably, PARP14 has gained increased attention as a promising target in cancer cells.