Fangchinoline targets human renal cell carcinoma cells through modulation of apoptotic and non­apoptotic cell deaths.

The process of apoptosis is one of the essential processes involved in maintenance of homeostasis in the human body. It can aid to remove misfolded proteins or cellular organelles. This sequence is especially necessary in cancer cells. However, specifically targeting already apoptotic pathways can induce drug resistance in cancer cells and hence drugs can induce cell death by alternative mechanism. We investigated whether fangchinoline (FCN) can target renal carcinoma cells by inducing multiple cell death mechanisms. Both paraptosis, autophagy, and apoptosis were induced by FCN through stimulation of diverse molecular signaling pathways. FCN induced ROS production with GSH/GSSG imbalance, and ER stress. In addition, formation of autophagosome and autophagy related markers were stimulated by FCN. Moreover, FCN induced cell cycle arrest and PARP cleavage. Except for blocking protein synthesis, these three cell death pathways were found to be complementarily working together with each other. FCN also exhibited synergistic effects with paclitaxel in inducing programmed cell death in RCC cells. Our data indicates that FCN could induce apoptotic cell death and non-apoptotic cell death pathways and can be con-tribute to development of novel cancer prevention or therapy.

Isoxazolyl-urea derivative evokes apoptosis and paraptosis by abrogating the Wnt/ß-catenin axis in colon cancer cells.

Deregulated activation of the Wnt/ß-catenin pathway is observed in many types of human malignancies including colon cancer. Abrogation of the Wnt/ß-catenin pathway has been demonstrated as an effective way of inducing cancer cell death. Herein, a new isoxazolyl-urea (QR-5) was synthesized and examined its efficacy on the viability of colon cancer cell lines. QR-5 displayed selective cytotoxicity towards colon cancer cells over normal counterparts. QR-5 induced apoptosis as evidenced by elevation in sub-G1 cells, decrease in Bcl-2, MMP-9, COX-2, VEGF and cleavage of PARP and caspase-3. QR-5 reduced the mitochondrial membrane potential, decreased the expression of Alix and elevated the expression of ATF4 and CHOP indicating the induction of paraptosis. The inhibitor of apoptosis (Z-DEVD-FMK) and paraptosis (CHX) could not restore Alix expression and PARP cleavage in QR-5 treated cells, respectively suggesting the complementation between the two cell death pathways. QR-5 suppressed the expression of Wnt/ß-catenin pathway proteins which was also evidenced by the downregulation of nuclear and cytoplasmic ß-catenin. The dependency of QR-5 on ß-catenin for inducing apoptosis and paraptosis was demonstrated by knockdown experiments using ß-catenin specific siRNA. Overall, QR-5 induces apoptosis as well as paraptosis by mitigating the Wnt/ß-catenin axis in colon cancer cells.

Kaempferide triggers apoptosis and paraptosis in pancreatic tumor cells by modulating the ROS production, SHP-1 expression, and the STAT3 pathway.

Pancreatic cancer is one of the deadliest diseases with a poor prognosis and a five-survival rate. The STAT3 pathway is hyperactivated which contributes to the sustained proliferative signals in pancreatic cancer cells. We have isolated kaempferide (KF), an O-methylated flavonol, from the green propolis of Mimosa tenuiflora and examined its effect on two forms of cell death namely, apoptosis and paraptosis. KF significantly increased the cleavage of caspase-3 and PARP. It also downmodulated the expression of Alix (an intracellular inhibitor of paraptosis) and increased the expression of CHOP and ATF4 (transcription factors that promote paraptosis) indicating that KF promotes apoptosis as well as paraptosis. KF also increased intracellular reactive oxygen species (ROS) suggesting the perturbance of the redox state. N-acetylcysteine reverted the apoptosis- and paraptosis-inducing effects of KF. Some ROS inducers are known to suppress the STAT3 pathway and investigation revealed that KF downmodulates STAT3 and its upstream kinases (JAK1, JAK2, and Src). Additionally, KF also elevated the expression of SHP-1, a tyrosine phosphatase which is involved in the negative modulation of the STAT3 pathway. Knockdown of SHP-1 prevented KF-driven STAT3 inhibition. Altogether, KF has been identified as a promoter of apoptosis and paraptosis in pancreatic cancer cells through the elevation of ROS generation and SHP-1 expression.

Brassinin alleviates cancer cachexia by suppressing diverse inflammatory mechanisms in mice.

Cancer cachexia is a multifactorial condition that contributes to the death of about 20% of cancer patients. It has the potential to cause weight loss, reduction in muscle mass, and loss of fat tissue, significantly lowering the quality of life. Currently, there are no approved drugs for cancer cachexia. Here, we have explored the possible impact of brassinin (BSN) on cancer cachexia under in vitro and in vivo settings. After differentiation, C2C12 and 3T3-L1 cells were incubated with colorectal carcinoma cells conditioned media or BSN. For preclinical studies, mice were injected with HT-29 cells followed by intraperitoneal administration of BSN, and muscle and adipose tissues were evaluated by Western blotting and hematoxylin and eosin staining. BSN effectively suppressed muscle atrophy by down-regulating the levels of Muscle RING-finger protein-1 and Atrogin-1, while also increasing the expression of myosin heavy chain in cachexia-induced-C2C12 myotubes. The induction of adipogenesis by BSN prevented adipocyte atrophy in cachexia-induced 3T3-L1 adipocytes. We also noted that BSN disrupted the interaction between COX-2 and signaling transducer and activator of transcription 3 (STAT3) promoter, leading to down-regulation of STAT3 activation. Moreover, it was found that BSN inhibited weight loss in mice and demonstrated anti-cachexic effects. Overall, our observations indicate that BSN can attenuate cancer cachexia through diverse mechanisms.

Unveiling autophagy complexity in leukemia: The molecular landscape and possible interactions with apoptosis and ferroptosis.

Autophagy is a self-digestion multistep process in which causes the homeostasis through degradation of macromolecules and damaged organelles. The autophagy-mediated tumor progression regulation has been a critical point in recent years, revealing the function of this process in reduction or acceleration of carcinogenesis. Leukemia is a haematological malignancy in which abnormal expansion of hematopoietic cells occurs. The current and conventional therapies from chemotherapy to cell transplantation have failed to appropriately treat the leukemia patients. Among the mechanisms dysregulated in leukemia, autophagy is a prominent one in which can regulate the hallmarks of this tumor. The protective autophagy inhibits apoptosis and ferroptosis in leukemia, while toxic autophagy accelerates cell death. The proliferation and invasion of tumor cells are tightly regulated by the autophagy. The direction of regulation depends on the function of autophagy that is protective or lethal. The protective autophagy accelerates chemoresistance and radio-resistsance. The non-coding RNAs, histone transferases and other pathways such as PI3K/Akt/mTOR are among the regulators of autophagy in leukemia progression. The pharmacological intervention for the inhibition or induction of autophagy by the compounds including sesamine, tanshinone IIA and other synthetic compounds can chance progression of leukemia.

Leonurine ameliorates the STAT3 pathway through the upregulation of SHP-1 to retard the growth of hepatocellular carcinoma cells.

Signal transducer and activator of transcription 3 (STAT3) is a transcription factor that directs the transcription of genes involved in the promotion of cell survival and proliferation, inflammation, angiogenesis, invasion, and migration. Overactivation of STAT3 is often witnessed in human cancers, thereby making it a good target in oncology. Herein the efficacy of Leonurine (Leo), a bioactive alkaloid present in Herba leonuri, was investigated for its STAT3-inhibitory potential in hepatocellular carcinoma (HCC) cells. Leo downregulated the persistent as well as IL-6-driven activation of STAT3. Leo abrogated the nuclear localization and DNA interacting ability of STAT3. Leo was also found to impart STAT3 inhibition by mitigating the activation of upstream kinases such as JAK1, JAK2, and Src both in constitutive and IL-6 inducible systems. Leo curbed the STAT3-driven luciferase gene expression and the depletion of STAT3 resulted in the reduced responsiveness of HCC cells to Leo. Pervanadate exposure counteracted Leo-induced STAT3 inhibition suggesting the involvement of a protein tyrosine phosphatase. SHP-1 was significantly elevated upon Leo exposure whereas the depletion of SHP-1 was found to revert the effect of Leo on STAT3. Leo induced apoptosis and also significantly potentiated the cytotoxic effect of paclitaxel, doxorubicin, and sorafenib. Leo was found to be non-toxic up to the dose of 10 mg/kg in NCr nude mice. In conclusion, Leo was demonstrated to induce cytotoxicity in HCC cells by mitigating the persistent of activation of STAT3 pathway.

CXCR4 expression is elevated in TNBC patient derived samples and Z-guggulsterone abrogates tumor progression by targeting CXCL12/CXCR4 signaling axis in preclinical breast cancer model.

Environmental factors such as exposure to ionizing radiations, certain environmental pollutants, and toxic chemicals are considered as risk factors in the development of breast cancer. Triple-negative breast cancer (TNBC) is a molecular variant of breast cancer that lacks therapeutic targets such as progesterone receptor, estrogen receptor, and human epidermal growth factor receptor-2 which makes the targeted therapy ineffective in TNBC patients. Therefore, identification of new therapeutic targets for the treatment of TNBC and the discovery of new therapeutic agents is the need of the hour. In this study, CXCR4 was found to be highly expressed in majority of breast cancer tissues and metastatic lymph nodes derived from TNBC patients. CXCR4 expression is positively correlated with breast cancer metastasis and poor prognosis of TNBC patients suggesting that suppression of CXCR4 expression could be a good strategy in the treatment of TNBC patients. Therefore, the effect of Z-guggulsterone (ZGA) on the expression of CXCR4 in TNBC cells was examined. ZGA downregulated protein and mRNA expression of CXCR4 in TNBC cells and proteasome inhibition or lysosomal stabilization had no effect on the ZGA-induced CXCR4 reduction. CXCR4 is under the transcriptional control of NF-κB, whereas ZGA was found to downregulate transcriptional activity of NF-κB. Functionally, ZGA downmodulated the CXCL12-driven migration/invasion in TNBC cells. Additionally, the effect of ZGA on growth of tumor was investigated in the orthotopic TNBC mice model. ZGA presented good inhibition of tumor growth and liver/lung metastasis in this model. Western blotting and immunohistochemical analysis indicated a reduction of CXCR4, NF-κB, and Ki67 in tumor tissues. Computational analysis suggested PXR agonism and FXR antagonism as targets of ZGA. In conclusion, CXCR4 was found to be overexpressed in majority of patient-derived TNBC tissues and ZGA abrogated the growth of TNBC tumors by partly targeting the CXCL12/CXCR4 signaling axis.

Methyl-Thiol-Bridged Oxadiazole and Triazole Heterocycles as Inhibitors of NF-κB in Chronic Myelogenous Leukemia Cells.

Nuclear factor kappa beta (NF-κB) is a transcriptional factor that plays a crucial role in regulating cancer cell proliferation. Therefore, the inhibition of NF-κB activity by small molecules may be beneficial in cancer therapy. In this report, methyl-thiol-bridged oxadiazole and triazole heterocycles were synthesized via click chemistry and it was observed that the lead structure, 2-(((1-(3,4-dichlorophenyl)-1H-1,2,3-triazol-4-yl)methyl)thio)-5-(4-methoxybenzyl)-1,3,4-oxadiazole (4c), reduced the viability of MCF-7 cells with an IC50 value of 7.4 µM. Compound 4c also caused concentration-dependent loss of cell viability in chronic myelogenous leukemia (CML) cells. Furthermore, compound 4c inhibited the activation of NF-κB in human CML cells as observed by nuclear translocation and DNA binding assays. Functionally, compound 4c produced PARP cleavage and also suppressed expression of Bcl-2/xl, MMP-9, COX-2, survivin, as well as VEGF, resulting in apoptosis of CML cells. Moreover, ChIP assay showed that compound 4c decreased the binding of COX-2 to the p65 gene promoter. Detailed in silico analysis also indicated that compound 4c targeted NF-κB in CML cells. In conclusion, a novel structure bearing both triazole and oxadiazole moieties has been identified that can target NF-κB in CML cells and may constitute a potential novel drug candidate.

Brucein D imparts a growth inhibitory effect in multiple myeloma cells by abrogating the Akt-driven signaling pathway.

The Akt signaling pathway is an oncogenic cascade activated in the bone marrow microenvironment of multiple myeloma (MM) cells and contributes to their uncontrolled proliferation. Abrogation of Akt signaling has been presented as one of the prime therapeutic targets in the treatment of MM. In the present report, we have investigated the effect of Brucein D (BD) on Akt-driven signaling events in MM cells. BD (300 nM) substantially inhibited cell viability and imparted growth-inhibitory effects in U266 cells as evidenced by cell viability assays and flow cytometric analysis. Effect of BD on cell viability was evaluated by MTT assay. Apoptotic cells and cell cycle arrest by BD were analyzed by flow cytometer. The results of the TUNEL assay and western blotting showed that BD induces apoptosis of MM cells by activating caspase-8 and 9 with subsequent reduction in the expression of antiapoptotic proteins (Bcl-2, Bcl-xl, survivin, cyclin D1, COX-2, VEGF, MMP-9). Analysis of activated kinases by Phospho-Kinase Array Kit revealed that Akt, p70S6K, HSP60, p53, and WNK1 were strongly expressed in untreated cells and BD treatment reversed this effect. Using transfection experiments, AKT depletion led to a decrease in phosphorylation of Akt, mTOR, p70S6K, and WNK. However, Akt overexpression led to increase in phosphorylation of these proteins. Depletion of Akt potentiated the apoptosis-inducing effect of BD whereas overexpression displayed resistance to BD-induced apoptosis suggesting the role of Akt in chemoresistance. Taken together, BD mitigates Akt-dependent signaling pathways in MM cells to impart its anticancer activity.

Potential Application of Leelamine as a Novel Regulator of Chemokine-Induced Epithelial-to-Mesenchymal Transition in Breast Cancer Cells.

CXCR7 and CXCR4 are G protein-coupled receptors (GPCRs) that can be stimulated by CXCL12 in various human cancers. CXCR7/4-CXCL12 binding can initiate activation of multiple pathways including JAK/STAT and manganese superoxide dismutase (MnSOD) signaling, and initiate epithelial-mesenchymal transition (EMT) process. It is established that cancer cell invasion and migration are caused because of these events. In particular, the EMT process is an important process that can determine the prognosis for cancer. Since the antitumor effect of leelamine (LEE) has been reported in various previous studies, here, we have evaluated the influence of LEE on the CXCR7/4 signaling axis and EMT processes. We first found that LEE suppressed expression of CXCR7 and CXCR4 both at the protein and mRNA levels, and showed inhibitory effects on these chemokines even after stimulation by CXCL12 ligand. In addition, LEE also reduced the level of MnSOD and inhibited the EMT process to attenuate the invasion and migration of breast cancer cells. In addition, phosphorylation of the JAK/STAT pathway, which acts down-stream of these chemokines, was also abrogated by LEE. It was also confirmed that LEE can induce an imbalance of GSH/GSSG and increases ROS, thereby resulting in antitumor activity. Thus, we establish that targeting CXCR7/4 in breast cancer cells can not only inhibit the invasion and migration of cancer cells but also can affect JAK/STAT, EMT process, and production of ROS. Overall, the findings suggest that LEE can function as a novel agent affecting the breast cancer.

Nimbolide enhances the antitumor effect of docetaxel via abrogation of the NF-κB signaling pathway in prostate cancer preclinical models.

Prostate cancer is the second most frequent type of cancer that affects men. Docetaxel (DTX) administration is the front-line therapy for patients with advanced prostate cancer and unfortunately, half of these patients develop resistance to DTX which could be due to its ability to activate the NF-κB pathway. The combinational effect of DTX and nimbolide on proliferation, apoptosis, activation of NF-κB, DNA binding ability of NF-κB, and expression of NF-κB-targeted gene products was investigated. The antitumor and antimetastatic effect of DTX or NL alone or in combination was also examined. The co-administration of NL and DTX resulted in a significant loss of cell viability with enhanced apoptosis in DTX-sensitive/resistant prostate cancer cells. NL abrogated DTX-triggered NF-κB activation and expression of its downstream antiapoptotic factors (survivin, Bcl-2, and XIAP). The combination of NL and DTX significantly reduced the DNA binding ability of NF-κB in both cell types. NL significantly enhanced the antitumor effect of DTX and reduced metastases in orthotopic models of prostate cancer. NL abolishes DTX-induced-NF-κB activation to counteract cell proliferation, tumor growth, and metastasis in the prostate cancer models.

Isoimperatorin down-regulates epithelial mesenchymal transition through modulating NF-κB signaling and CXCR4 expression in colorectal and hepatocellular carcinoma cells.

Worldwide, metastatic spread of tumor is the major cause of cancer-related deaths. There are many anti-metastatic agents that can target metastasis-related pathways, but there are relatively few studies on agents targeting C-X-C chemokine receptor type 4 (CXCR4) signaling axis. Here, we have investigated whether Isoimperatorin (IIPT), derived from Angelica dahurica can modulate CXCR4 signaling axis-related epithelial-to-mesenchymal transition (EMT) and tumor metastasis process. We evaluated the influence of IIPT on CXCR4, HER2, MMPs, EMT markers, and NF-κB signaling pathway by using Western blot analysis. The cellular invasion and migration were observed by Boyden chamber and wound healing assays. IIPT has a down-regulatory effect on CXCR4, HER2, and MMP-9/2. On the contrary, imperatorin (IPT) as compared to IIPT did not alter the expression of CXCR4. IIPT down-regulated the protein levels and RNA levels of mesenchymal markers, twist, snail, and enhanced the levels of different epithelial markers. IIPT also inhibited cell migration, invasion, and proliferation. Furthermore, IIPT negatively regulated constitutive NF-κB activation and inhibited the translocation of phospho-p65 and p65 into the nuclei. IIPT can potentially function as a novel anti-metastatic agent by inhibiting EMT and metastasis process via inhibition of NF-κB activation and CXCR4 expression in colorectal and hepatocellular carcinoma cells.

2,3,5,6-Tetramethylpyrazine Targets Epithelial-Mesenchymal Transition by Abrogating Manganese Superoxide Dismutase Expression and TGFß-Driven Signaling Cascades in Colon Cancer Cells.

Epithelial-mesenchymal transition (EMT) is a crucial process in which the polarized epithelial cells acquire the properties of mesenchymal cells and gain invasive properties. We have previously demonstrated that manganese superoxide dismutase (MnSOD) can regulate the EMT phenotype by modulating the intracellular reactive oxygen species. In this report, we have demonstrated the EMT-suppressive effects of 2,3,5,6-Tetramethylpyrazine (TMP, an alkaloid isolated from Chuanxiong) in colon cancer cells. TMP suppressed the expression of MnSOD, fibronectin, vimentin, MMP-9, and N-cadherin with a parallel elevation of occludin and E-cadherin in unstimulated and TGFß-stimulated cells. Functionally, TMP treatment reduced the proliferation, migration, and invasion of colon cancer cells. TMP treatment also modulated constitutive activated as well as TGFß-stimulated PI3K/Akt/mTOR, Wnt/GSK3/ß-catenin, and MAPK signaling pathways. TMP also inhibited the EMT program in the colon cancer cells-transfected with pcDNA3-MnSOD through modulation of MnSOD, EMT-related proteins, and oncogenic pathways. Overall, these data indicated that TMP may inhibit the EMT program through MnSOD-mediated abrogation of multiple signaling events in colon cancer cells.

Fangchinoline abrogates growth and survival of hepatocellular carcinoma by negative regulation of c-met/HGF and its associated downstream signaling pathways.

Among all cancers, hepatocellular carcinoma (HCC) remains a lethal disease with limited treatment options. In this study, we have analyzed the possible inhibitory effects of Fangchinoline (FCN) on c-Met, a protein known to regulate the rapid phosphorylation of downstream signals, as well as mediate aberrant growth, metastasis, survival, and motility in cancer. FCN inhibited the activation of c-Met and its downstream signals PI3K, AKT, mTOR, MEK, and ERK under in vitro settings. Moreover, c-Met gene silencing lead to suppression of PI3K/AKT/mTOR and MEK/ERK signaling pathways, and induced apoptotic cell death upon exposure to FCN. In addition, FCN markedly inhibited the expression of the various oncogenic proteins such as Bcl-2/xl, survivin, IAP-1/2, cyclin D1, and COX-2. In vivo studies in HepG2 cells xenograft mouse model showed that FCN could significantly attenuate the tumor volume and weight, without affecting significant loss in the body weight. Similar to in vitro studies, expression level of c-Met and PI3K/AKT/mTOR, MEK/ERK signals was also suppressed by FCN in the tissues obtained from mice. Therefore, the novel findings of this study suggest that FCN can potentially function as a potent anticancer agent against HCC.

Tanshinone IIA exerts autophagic cell death through down-regulation of ß-catenin in renal cell carcinoma cells.

Renal cell carcinoma (RCC), also called kidney cancer, is one of the most common malignancies worldwide, including the United States and China. Because of the characteristics of RCC that are both insidious and largely insensitive to chemo-radiation, the incidence and mortality of RCC are increasing every year. However, there are few studies describing anti-cancer effects of the natural compounds on RCC as compared to other cancers. Here, we analyzed the anti-neoplastic impact of Tanshinone IIA (TSN) on RCC cells. We noted that TSN increased the expression of LC3 proteins while having little effect on PARP and Alix protein expression. We found that TSN up-regulated the expression of autophagy-related proteins such as Atg7 and Beclin-1. Moreover, TSN promoted the formation of autophagic vacuoles such as autophagosomes and autolysosomes. However, treatment with 3-Methyladenine (3-MA) or Chloroquine (CQ), slightly decreased the ability of TSN to induce autophagy, but still autophagy occurred. In addition, TSN inhibited translocation of ß-catenin into the nucleus, and ß-catenin deletion and TSN treatment in RCC increased the expression of LC3 protein. Overall, our findings indicate that TSN can exert significant anti-tumor effects through down-regulation of ß-catenin to induce autophagic cell death.

Withanolide modulates the potential crosstalk between apoptosis and autophagy in different colorectal cancer cell lines.

Withaferin A (WFA), a withanolide, is isolated from plants of Withania somnifera (L.) Dual (Solanaceae), known as Indian ginseng, Indian winter cherry or Ashwagandha. It has been reported to exert multifaceted anti-neoplastic effects. Here, we analyzed the impact of WFA on apoptosis and autophagy activation in different human colorectal cancer cell lines. We observed that WFA exposure caused an increased aggregation of cells in the subG1 arrest in cell cycle, and increased the number of late apoptotic cells. WFA also induced the apoptosis via PARP and caspase-3 cleavage accompanied with suppression of levels of anti-apoptotic proteins like Bcl-2 and Bcl-xl. The influence of WFA on autophagy was validated by acridine orange, MDC staining, and immunocytochemistry of LC3. It was found that 24 h treatment of WFA increased the acridine and MDC stained autophagosome with induced the LC3 and other autophagy markers Atg7 and beclin-1 activation. We used Z-DEVD-FMK, a caspase-3 blocker, and 3-MA, an autophagy inhibitor, to confirm whether these effects were specific to apoptosis and autophagy, and observed the recovery of both these processes upon exposure to WFA. Moreover, the activation of ß-catenin protein was attenuated by WFA. Interestingly, small interfering RNA (siRNA)-promoted ß-catenin knockdown augmented the WFA-induced active form of p-GSK-3ß, and stimulated autophagy and apoptosis through PARP and LC3 activation. These findings suggested that WFA could stimulate activation of both apoptosis and autophagy process via modulating ß-catenin pathway.

Regulation of apoptosis and autophagy by albendazole in human colon adenocarcinoma cells.

Albendazole (ABZ) was initially introduced as an anthelmintic, however, many studies have reported with its anticancer effects. We investigated the anti-tumor effects of ABZ in vitro in human colon adenocarcinoma HCT-15, HCT-116, HT-29, and SW480 cell lines in this study. The cytotoxicity of ABZ was analyzed in colon adenocarcinoma cell lines and normal CCD18Co cells. We found that ABZ induced the subG1 arrest during cell cycle progression, increased the late apoptotic cells, shifted of peak TUNEL-labeled cells peak, and induced apoptosis. Then effects on autophagy activation was confirmed by acridine orange (AO), MDC staining, and immunocytochemistry of LC3. It was observed that ABZ can induce the autophagy activation through modulating the levels of LC3, Atg7, and beclin-1. For mechanistic studies, apoptosis blocker (Z-DEVD-FMK) and autophagy inhibitor (3-MA) were used to confirm that whether ABZ has apoptosis and autophagy specific effects, and reversal in both these cell death processes were noted. The effects of ABZ on AMPK, MAPKs, and ULK induction was also evaluated. We noticed that N-acetyl cysteine (NAC), a broad spectrum antioxidant, can effectively inhibit both apoptosis and autophagy. However, ABZ could even recover suppression of apoptosis and autophagy caused by NAC in colon cancer cells. Therefore, ABZ can potentially up-regulate both the apoptosis and autophagy to significantly suppress tumorigenesis in colorectal cancer cell lines.

Leelamine Exerts Antineoplastic Effects in Association with Modulating Mitogen­Activated Protein Kinase Signaling Cascade.

Mitogen­activated protein kinase (MAPK) pathway is a prominent signaling cascade that modulates cell proliferation, apoptosis, stress response, drug resistance, immune response, and cell motility. Activation of MAPK by various small molecules/natural compounds has been demonstrated to induce apoptosis in cancer cells. Herein, the effect of leelamine (LEE, a triterpene derived from bark of pine trees) on the activation of MAPK in hepatocellular carcinoma (HCC) and breast cancer (BC) cells was investigated. LEE induced potent cytotoxicity of HCC (HepG2 and HCCLM3) and BC (MDA-MB-231 and MCF7) cells over normal counterparts (MCF10A). LEE significantly enhanced the phosphorylation of p38 and JNK MAPKs in a dose-dependent fashion and it did not affect the phosphorylation of ERK in HCC and BC cells. The apoptosis-driving effect of LEE was further demonstrated by cleavage of procaspase-3/Bid and suppression of prosurvival proteins (Bcl-xL and XIAP). Furthermore, LEE also reduced the SDF1-induced-migration and -invasion of HCC and BC cells. Taken together, the data demonstrated that LEE promotes apoptosis and induces an anti-motility effect by activating p38 and JNK MAPKs in HCC and BC cells.

Leelamine Modulates STAT5 Pathway Causing Both Autophagy and Apoptosis in Chronic Myelogenous Leukemia Cells.

Leelamine (LEE) has recently attracted significant attention for its growth inhibitory effects against melanoma, breast cancer, and prostate cancer cells; however, its impact on hematological malignancies remains unclear. Here, we first investigate the cytotoxic effects of LEE on several human chronic myeloid leukemia (CML) cells. We noted that LEE stimulated both apoptosis and autophagy in CML cells. In addition, the constitutive activation of signal transducer and activator of transcription 5 (STAT5) was suppressed substantially upon LEE treatment. Moreover, STAT5 knockdown with small interfering RNA (siRNA) increased LEE-induced apoptosis as well as autophagy and affected the levels of various oncogenic proteins. Thus, the targeted mitigation of STAT5 activation by LEE can contribute to its diverse anticancer effects by enhancing two distinct cell death pathways.

Fangchinoline diminishes STAT3 activation by stimulating oxidative stress and targeting SHP-1 protein in multiple myeloma model.

Introduction: The development of cancer generally occurs as a result of various deregulated molecular mechanisms affecting the genes that can control normal cellular growth. Signal transducer and activator of transcription 3 (STAT3) pathway, once aberrantly activated can promote carcinogenesis by regulating the transcription of a number of oncogenic genes. Objectives: Here, we evaluated the impact of fangchinoline (FCN) to attenuate tumor growth and survival through modulation of oncogenic STAT3 signaling pathway using diverse tumor cell lines and a xenograft mouse model. Methods: To evaluate the action of FCN on STAT3 cascade, protein levels were analyzed by Western blot analysis and electrophoretic mobility shift assay (EMSA). Translocation of STAT3 was detected by immunocytochemistry. Thereafter, FCN-induced ROS was measured by GSH/GSSG assay and H2DCF-DA. FCN-induced apoptosis was analyzed using Western blot analysis and flow cytometry for various assays. Finally, anti-cancer effects of FCN in vivo was evaluated in a myeloma model. Results: We noted that FCN abrogated protein expression levels of STAT3 and upstream signals (JAK1/2 and Src). In addition, FCN also attenuated DNA binding ability of STAT3 and its translocation into the nucleus. It altered the levels of upstream signaling proteins, increased SHP-1 levels, and induced substantial apoptosis in U266 cells. FCN also promoted an increased production of reactive oxygen species (ROS) and altered GSSG/GSH ratio in tumor cells. Moreover, FCN effectively abrogated tumor progression and STAT3 activation in a preclinical myeloma model. Conclusion: Overall, this study suggests that FCN may have a tremendous potential to alter abnormal STAT3 activation and induce cell death in malignant cells along with causing the suppression of pathogenesis and growth of cancer through a pro-oxidant dependent molecular mechanism.