Post-marketing safety of immunomodulatory drugs in multiple myeloma: A pharmacovigilance investigation based on the FDA adverse event reporting system.

Objective: In recent years, the emergence of immunomodulatory drugs (IMiDs) has significantly improved clinical outcomes in patients with multiple myeloma (MM); however, serious adverse events (AEs) have hindered their safe clinical application. This study aimed to characterize the safety profiles and differences in IMiDs through a disproportionality analysis using the U.S. Food and Drug Administration Adverse Event Reporting System (FAERS), a post-marketing surveillance database. Methods: This study filtered reports of thalidomide, lenalidomide, and pomalidomide as primary suspect drugs in FAERS files from January 2013 to December 2021. AEs in the reports were retrieved according to the preferred terms (PTs) of the Medical Dictionary for Regulatory Activities. Furthermore, we detected safety signals using the reporting odds ratio (ROR), proportional reporting ratio (PRR), and Bayesian belief propagation neural network (BCPNN). When all three algorithms showed an association between the target drug and the AE, a positive signal was generated. Results: We extracted 9,968 thalidomide, 231,926 lenalidomide, and 55,066 pomalidomide AE reports. AEs were more common in male patients and in those >44 years old. Important safety signals were detected based on the system organ classes (SOC), including thalidomide (cardiac disorders: ROR, 2.87; PRR, 2.79; IC 1.22), lenalidomide (gastrointestinal disorders: ROR, 2.38; PRR, 2.27; IC 0.75), and pomalidomide (respiratory, thoracic, and mediastinal disorders: ROR, 2.14; PRR, 2.09; IC 0.85). Within the PT level, we identified novel risk signals: the thalidomide-induced second primary malignancy (SPM) signal was significant; lenalidomide reduced the success rate of hematopoietic stem cell collection; and three IMiDs may cause human chorionic gonadotropin increase, but this needs to be proven by clinical data. Pneumonia, sepsis, and renal failure are common risk factors for death due to IMiDs. Compared with thalidomide and lenalidomide, pomalidomide has a lower risk of venous thromboembolism (VTE) and is beneficial to patients with renal insufficiency. Conclusion: Mining data from FAERS resulted in novel AE signals, including adenocarcinoma of colon, harvest failure of blood stem cells, and increased levels of human chorionic gonadotropin. Further investigation is required to verify the significance of these signals. Moreover, IMiDs showed differences in safety reports, which should be emphasized by clinicians.

The pathogenesis, diagnosis, prevention, and treatment of CAR-T cell therapy-related adverse reactions.

Chimeric antigen receptor (CAR)-T cell therapy is effective in the treatment of refractory/relapsed (r/r) hematological malignancies (r/r B-cell lymphoblastic leukemia, B-cell lymphoma, and multiple myeloma). In addition, it is being explored as a treatment option for solid tumors. As of 31 March 2022, seven CAR-T therapies for hematological malignancies have been approved worldwide. Although CAR-T therapy is an effective treatment for many malignancies, it also causes adverse effects. The incidence of cytokine release syndrome (CRS), the most common adverse reaction after infusion of CAR-T cells, is as high as 93%.CRS, is the leading risk factor of immune effector cell-associated neurotoxicity syndrome (ICANS), as well as cardiovascular, hematological, hepatorenal, skin, pulmonary, and gastrointestinal toxicity. Severe adverse reactions complicated by CRS severely impede the widespread application of CAR-T therapy. The CAR-T product was initially approved in 2017; however, only limited studies have investigated the adverse reactions owing to CAR-T therapy compared to that of clinically approved drugs. Thus, we aimed to elucidate the mechanisms, risk factors, diagnostic criteria, and treatment of toxicities concurrent with CRS, thereby providing a valuable reference for the safe, effective, and widespread application of CAR-T therapy.

Mechanism of Lethal Skin Toxicities Induced by Epidermal Growth Factor Receptor Inhibitors and Related Treatment Strategies.

Epidermal growth factor receptor (EGFR) inhibitors are widely used to treat various types of cancers such as non-small cell lung cancer, head and neck cancer, breast cancer, pancreatic cancer. Adverse reactions such as skin toxicity, interstitial lung disease, hepatotoxicity, ocular toxicity, hypomagnesemia, stomatitis, and diarrhea may occur during treatment. Because the EGFR signaling pathway is important for maintaining normal physiological skin function. Adverse skin reactions occurred in up to 90% of cancer patients treated with EGFR inhibitors, including common skin toxicities (such as papulopustular exanthemas, paronychia, hair changes) and rare fatal skin toxicities (e.g., Stevens-Johnson syndrome, toxic epidermal necrolysis, acute generalized exanthematous pustulosis). This has led to the dose reduction or discontinuation of EGFR inhibitors in the treatment of cancer. Recently, progress has been made about research on the skin toxicity of EGFR inhibitors. Here, we summarize the mechanism of skin toxicity caused by EGFR inhibitors, measures to prevent severe fatal skin toxicity, and provide reference for medical staff how to give care and treatment after adverse skin reactions.

Neuroprotective Effects of Tetrahydroxystilbene Glucoside against Rotenone-Induced Toxicity in PC12 Cells.

To investigate the mechanism of the protective effect of tetrahydroxystilbene glucoside (TSG) on nerve cells, an injury model induced by rotenone in PC12 cells was constructed. Cell viability was detected by using cell counting kit-8 (CCK8) assay. Apoptosis was detected by using flow cytometry. The mitochondrial membrane potential (MMP) was detected by using the fluorescent probe JC-1. Generation of reactive oxygen species (ROS) in PC12 cells was determined using the 5-(and-6)-chloromethyl-2',7'-dichlorodihydrofluorescein diacetate acetyl ester (CM-H2DCFDA) probe. Protein expression in PC12 cells was detected using Western blotting. The results showed that TSG (20-100 µM) attenuated the cytotoxic effects of rotenone on PC12 cells. TSG pretreatment attenuated the apoptosis rate, the degradation of poly(ADP-ribose)polymerase (PARP) and the activation of cleaved caspase 3, which was induced by rotenone. TSG can significantly reduce the effect of rotenone on the reduction of MMP and the expression of cytoC in the cytosolic fraction. TSG attenuated rotenone-induced de-phosphorylation and mitochondrial translocation of cofilin, as well as rotenone-induced accumulation of ROS. The Western blot results showed that ROT could decrease the expression level of phosphorylated (p)-Glycogen synthase kinase-3ß (GSK)-3ß and p-AKT, and TSG could weaken these effects of rotenone. In addition, TSG increased the expression level of nuclear factor-E2-related factor 2 (Nrf2) in the nuclear fraction. These results suggest that TSG could protect PC12 cells against rotenone through multiple pathways. Thus, TSG has the potential to become a novel neuroprotective agent.

The Application of Peptides in Glioma: A Novel Tool for Therapy.

BACKGROUND: Glioma is the most aggressive and lethal tumor of the central nervous system. Due to the cellular heterogeneity, the invasiveness, and blood-brain barrier (BBB), current therapeutic approaches, such as chemotherapy and radiotherapy, are poorly to obtain great antitumor efficacy. However, peptides, a novel type of therapeutic agent, displayed excellent ability in the tumor, which becomes a new molecule for glioma treatment. METHODS: We review the current knowledge on peptides for the treatment of glioma through a PubMed-based literature search. RESULTS: In the treatment of glioma, peptides can be used as (i) decoration on the surface of the delivery system, facilitating the distribution and accumulation of the anti-tumor drug in target site; (ii) anti-tumor active molecules, inhibiting the growth of glioma and reducing solid tumor volume; (iii) immune-stimulating factor, and it activating immune cells in the tumor microenvironment or recruiting immune cells to the tumor for breaking out the immunosuppression by glioma cells. CONCLUSION: The application of peptides has revolutionized the treatment of glioma, which based on targeting, penetrating, anti-tumor activities and immunostimulatory. Moreover, better outcomes have been discovered in combining different kinds of peptides rather than a single one. Until now, more and more preclinical studies have been developed with multifarious peptides, which shows promising results in vitro or vivo with the model of glioma.

Cyclovirobuxine D Induces Apoptosis and Mitochondrial Damage in Glioblastoma Cells Through ROS-Mediated Mitochondrial Translocation of Cofilin.

BACKGROUND: Cyclovirobuxine D (CVBD), a steroidal alkaloid, has multiple pharmacological activities, including anti-cancer activity. However, the anti-cancer effect of CVBD on glioblastoma (GBM) has seldom been investigated. This study explores the activity of CVBD in inducing apoptosis of GBM cells, and examines the related mechanism in depth. METHODS: GBM cell lines (T98G, U251) and normal human astrocytes (HA) were treated with CVBD. Cell viability was examined by CCK-8 assay, and cell proliferation was evaluated by cell colony formation counts. Apoptosis and mitochondrial superoxide were measured by flow cytometry. All protein expression levels were determined by Western blotting. JC-1 and CM-H2DCFDA probes were used to evaluate the mitochondrial membrane potential (MMP) change and intracellular ROS generation, respectively. The cell ultrastructure was observed by transmission electron microscope (TEM). Colocalization of cofilin and mitochondria were determined by immunofluorescence assay. RESULTS: CVBD showed a greater anti-proliferation effect on the GBM cell lines, T98G and U251, than normal human astrocytes in dose- and time-dependent manners. CVBD induced apoptosis and mitochondrial damage in GBM cells. We found that CVBD led to mitochondrial translocation of cofilin. Knockdown of cofilin attenuated CVBD-induced apoptosis and mitochondrial damage. Additionally, the generation of ROS and mitochondrial superoxide was also induced by CVBD in a dose-dependent manner. N-acetyl-L-cysteine (NAC) and mitoquinone (MitoQ) pre-treatment reverted CVBD-induced apoptosis and mitochondrial damage. MitoQ pretreatment was able to block the mitochondrial translocation of cofilin caused by CVBD. CONCLUSIONS: Our data revealed that CVBD induced apoptosis and mitochondrial damage in GBM cells. The underlying mechanism is related to mitochondrial translocation of cofilin caused by mitochondrial oxidant stress.

Bovine Serum Albumin Nanoparticle-Mediated Delivery of Sorafenib for Improving Hepatocellular Carcinoma Therapy.

Hepatocellular carcinoma (HCC) is the most common primary malignancy of the liver. The majority of patients with HCC are diagnosed with advanced-stage disease. Sorafenib is a frontline therapy drug approved by the Food and Drug Administration for advanced HCC. However, the poor aqueous solubility of sorafenib limits its applications. The present study aimed to overcome this limitation of sorafenib. Thus, bovine serum albumin (BSA)-based nanoparticles were developed to encapsulate hydrophobic sorafenib. The resultant sorafenib-loaded BSA nanoparticles (Sf-BSA-NPs) were thoroughly characterized for size distribution, encapsulation efficiency and morphology. Previous studies on HepG2 cells in vitro have demonstrated that Sf-BSA-NPs exhibit remarkable superiority to free sorafenib in cytocompatibility, cytotoxicity and proapoptotic effect. The results of the present study demonstrated that Sf-BSA-NPs were effective in improving aqueous solubility, and enhanced drug cytotoxicity, suggesting its therapeutic potential for HCC.

The Immunostimulative Effect and Mechanisms of a Novel Mouse Anti-Human PD-1 Monoclonal Antibody on Jurkat Lymphocytic Cells Cocultured with Hepatoma Cells.

BACKGROUND: Monoclonal antibodies (mAbs) that target the programmed cell death-1 (PD-1)/programmed death-ligand 1 (PD-L1) immune checkpoint have demonstrated substantial clinical benefit for a variety of solid tumors. However, their applications in patients with hepatocellular carcinoma (HCC) are reported with unclear molecular mechanisms. Here, we report a novel mouse anti-human PD-1 mAb that can reverse the immunosuppressive effect of HePG2 cells on Jurkat cells. MATERIALS AND METHODS: HepG2 liver cancer cells, which were induced to overexpress PD-L1 by IFN-γ, were co-cultured with PHA-activated Jurkat lymphocytic cells to investigate the immunostimulative effect and mechanisms of the 14 newly generated PD-1 mAbs. Multiple cellular and molecular biology experiments were performed in this study, such as CCK-8, ELISA, flow cytometry, immunofluorescence and Western blot. RESULTS: We found that mAb B1C4 significantly enhanced the tumor-killing cytokine secretion level by Jurkat cells in the co-culture system and increased the killing ability of Jurkat cells on HepG2 cells. Co-culture with HePG2 cells led to Jurkat cell cycle delay in S phase, and B1C4 promoted cell cycle progression from S to G2/M. Co-culture with HePG2 cells also caused apoptosis in Jurkat cells, which was inhibited by B1C4. B1C4 reversed the immunosuppression of Jurkat cells resulted from co-cultured with HePG2 cells through inhibiting PTEN and activating PI3K/AKT/mTOR signaling pathways. CONCLUSION: Our study demonstrated that anti-PD-1 mAb B1C4 could inhibit the apoptosis of Jurkat cells induced by HePG2 hepatoma cells and reverse the immunosuppressive effect of HePG2 cells on Jurkat cells. The study provides a vital basis for applying PD-1 monoclonal antibodies in the treatment of HCC and provides antibody selection for the development of novel PD-1 mAb with blocking activity.

Hyperoside Protected Against Oxidative Stress-Induced Liver Injury via the PHLPP2-AKT-GSK-3ß Signaling Pathway In Vivo and In Vitro.

Hyperoside, isolated from Drosera rotundifolia L., seeds of Cuscuta chinensis Lam., or Hypericum perforatum L., originally showed to possess an antifungal and antibacterial activity, while recently showed the protective effects against oxidative stress-induced liver injury. This study investigated such a protective effect of hyperoside and the underlying molecular mechanisms in vitro and in carbon tetrachloride (CCl4)-injured rat livers. The data showed that hyperoside was able to prevent the oxidative stress-induced liver morphological changes and CCl4-induced rat liver injury. Hyperoside reversed the decrease of superoxidase dismutase (SOD) level and the increase of malondialdehyde (MDA) level in vivo. Moreover, hyperoside regulated the pleckstrin homology (PH) domain leucine-rich repeat protein phosphatase 2 (PHLPP2)-protein kinase B (AKT)-glycogen synthase kinase 3ß (GSK-3ß) signaling pathway in tert-butylhydroquinone (t-BHP)-treated liver cells, e.g., Hyperoside reduced PHLPP2 expression to activate AKT phosphorylation, induce GSK-3ß phosphorylation, and then increased nuclear factor erythroid-2-related factor 2 (Nrf2) nuclear translocation, reduced nuclear translocation of phosphorylated Fyn, and promoted heme oxygenase-1 (HO-1) expression in vivo and in vitro. In contrast, siRNA-mediated knockdown of PHLPP2 expression enhanced hyperoside-mediated activation of the AKT-GSK-3ß kinase pathway in liver cells. In conclusion, the present study demonstrated that hyperoside could protect against oxidative stress-induced liver injury by regulating the PHLPP2-AKT-GSK-3ß signaling pathway in vivo and in vitro.

Recent Developments in Pharmacological Effect, Mechanism and Application Prospect of Diazeniumdiolates.

Nitric oxide (NO) is a simple structured and unstable free radical molecule, which participates in the regulation of many pathophysiological processes. It functions both as a second messenger and as an endogenous neurotransmitter. Diazeniumdiolates (NONOates) are a series of compounds containing the functional parent nuclear structure of [N(O)NO]-, which are the most widely studied NO donors. NONOates are unstable and easy to release NO in physiological conditions. The biomedical applications and drug development of NO donor have attracted the scientists' attention in recent years. In this review, recent advances in NONOates research are highlighted in terms of chemical structures, molecular characteristics, pharmacological effects, and biomedical application prospects.

Combination of Detoxified Pneumolysin Derivative ΔA146Ply and Berbamine as a Treatment Approach for Breast Cancer.

Increasing evidence demonstrates that microorganisms and their products can modulate host responses to cancer therapies and contribute to tumor shrinkage via various mechanisms, including intracellular signaling pathways modulation and immunomodulation. Detoxified pneumolysin derivative ΔA146Ply is a pneumolysin mutant lacking hemolytic activity. To determine the antitumor activity of ΔA146Ply, the combination of ΔA146Ply and berbamine, a well-established antitumor agent, was used for breast cancer therapy, especially for triple-negative breast cancer. The efficacy of the combination therapy was evaluated in vitro using four breast cancer cell lines and in vivo using a synergistic mouse tumor model. We demonstrated that in vitro, the combination therapy significantly inhibited cancer cell proliferation, promoted cancer cell apoptosis, caused cancer cell-cycle arrest, and suppressed cancer cell migration and invasion. In vivo, the combination therapy significantly suppressed tumor growth and prolonged the median survival time of tumor-bearing mice partially through inhibiting tumor cell proliferation, promoting tumor cell apoptosis, and activating systemic antitumor immune responses. The safety analysis demonstrated that the combination therapy showed no obvious liver and kidney toxicity to tumor-bearing mice. Our study provides a new treatment option for breast cancer and lays the experimental basis for the development of ΔA146Ply as an antitumor agent.

Saikosaponin D inhibits autophagosome­lysosome fusion and induces autophagy­independent apoptosis in MDA­MB­231 breast cancer cells.

The present study aimed to explore the effect of Saikosaponin D (SSD) and its underlying mechanism on apoptosis and autophagy in human breast cancer MDA­MB­231 cells. MTT assay, flow cytometry, western blotting and confocal fluorescence microscopy detection were employed. SSD, a kind of triterpenoid saponins extracted from Radix bupleuri, has been demonstrated to have the effects of anti­inflammatory, antioxidative and anticancer effects and can regulate autophagy. The present study revealed that SSD induced apoptosis through the activation of the p38 mitogen­activated protein kinase (MAPK) signaling pathway in human breast cancer MDA­MB­231 cells. The administration of SSD promoted the phosphorylation/activation of p38 MAPK in MDA­MB­231 cells, whereas pretreatment with SB203580, an effective p38 MAPK inhibitor, attenuated SSD­mediated apoptosis, the cleavage of PARP and the activation of caspase­3. In addition, SSD blocked autophagic degradation by inhibiting autolysosome formation, resulting in the accumulation of autophagosomes. Mechanistically, the results of the present study revealed that SSD inhibited the formation of autophagosomes by inhibiting autophagosome­lysosome fusion, rather than by damaging lysosome function. Furthermore, blocking autophagy degradation was not associated with SSD­mediated apoptosis. The genetic knockdown of autophagy­related protein 5 markedly reduced SSD­mediated LC3B­II accumulation; however, it did not affect the SSD­mediated phosphorylation/activation of p38, cleavage of PARP, activation of caspase­3 or apoptosis. In conclusion, the findings of the present study suggest that SSD may induce apoptosis and block autophagic degradation, which provides further evidence of the association between the inhibition of autophagic degradation and cell death.

Downregulation of MYO1C mediated by cepharanthine inhibits autophagosome-lysosome fusion through blockade of the F-actin network.

BACKGROUND: MYO1C, an actin-based motor protein, is involved in the late stages of autophagosome maturation and fusion with the lysosome. The molecular mechanism by which MYO1C regulates autophagosome-lysosome fusion remains largely unclear. METHODS: Western blotting was used to determine the expression of autophagy-related proteins. Transmission electron microscopy (TEM) was used to observe the ultrastructural changes. An immunoprecipitation assay was utilized to detect protein-protein interactions. Immunofluorescence analysis was used to detect autophagosome-lysosome fusion and colocalization of autophagy-related molecules. An overexpression plasmid or siRNA against MYO1C were sequentially introduced into human breast cancer MDA-MB-231 cells. RESULTS: We show here that cepharanthine (CEP), a novel autophagy inhibitor, inhibited autophagy/mitophagy through blockage of autophagosome-lysosome fusion in human breast cancer cells. Mechanistically, we found for the first time that MYO1C was downregulated by CEP treatment. Furthermore, the interaction/colocalization of MYO1C and F-actin with either LC3 or LAMP1 was inhibited by CEP treatment. Knockdown of MYO1C further decreased the interaction/colocalization of MYO1C and F-actin with either LC3 or LAMP1 inhibited by CEP treatment, leading to blockade of autophagosome-lysosome fusion. In contrast, overexpression of MYO1C significantly restored the interaction/colocalization of MYO1C and F-actin with either LC3 or LAMP1 inhibited by CEP treatment. CONCLUSION: These findings highlight a key role of MYO1C in the regulation of autophagosome-lysosome fusion through F-actin remodeling. Our findings also suggest that CEP could potentially be further developed as a novel autophagy/mitophagy inhibitor, and a combination of CEP with classic chemotherapeutic drugs could become a promising treatment for breast cancer.

ROS-mediated activation and mitochondrial translocation of CaMKII contributes to Drp1-dependent mitochondrial fission and apoptosis in triple-negative breast cancer cells by isorhamnetin and chloroquine.

BACKGROUND: Triple-negative breast cancer (TNBC) is often aggressive and associated with a poor prognosis. Due to the lack of available targeted therapies and to problems of resistance with conventional chemotherapeutic agents, finding new treatments for TNBC remains a challenge and a better therapeutic strategy is urgently required. METHODS: TNBC cells and xenograft mice were treated with a combination of chloroquine (CQ) and isorhamnetin (IH). Mitochondrial fission, apoptosis, and related signaling pathways were determined by flow cytometry, immunofluorescence, and related molecular biological techniques. RESULTS: The inhibition of autophagy/mitophagy by CQ selectively enhances IH-induced mitochondrial fission and apoptosis in TNBC cells but not in estrogen-dependent breast cancer cells. These events were accompanied by mitochondrial translocation of Bax and the release of cytochrome c. Mechanistically, these effects were associated with oxidative stress-mediated phosphorylation of CaMKII (Thr286) and Drp1 (S616), and subsequent mitochondrial translocation of CaMKII and Drp1. The interruption of the CaMKII pathway by genetic approaches (e.g. CaMKII mutant or siRNA) attenuated combination-mediated mitochondrial fission and apoptosis. The combination of CQ/IH was a marked inhibitor tumor growth, inducing apoptosis in the TNBC xenograft mouse model in association with the activation of CaMKII and Drp1 (S616). CONCLUSIONS: Our study highlights the critical role of ROS-mediating CaMKII/Drp1 signaling in the regulation of mitochondrial fission and apoptosis induced by combination of CQ/IH. These findings also suggest that IH could potentially be further developed as a novel chemotherapeutic agent. Furthermore, a combination of IH with classic autophagy/mitophagy inhibitor could represent a novel therapeutic strategy for the treatment of TNBC.

Ars2 promotes cell proliferation and tumorigenicity in glioblastoma through regulating miR-6798-3p.

Arsenic resistance protein 2 (Ars2) is a component of the nuclear RNA cap-binding complex (CBC) that is important for some microRNA biogenesis and it is critical for cell proliferation and tumorigenicity. However, mechanism of Ars2-regulated cellular proliferation and tumorigenicity in glioblastoma has not been fully understood. Western blotting was used to detect the expressions of Ars2, p53, p21, and cleavage/activation of caspases-3 (C-Caspase 3). Microarray and Quantitative Real-time PCR (qRT-PCR) were performed to identify the Ars2-regulated microRNAs. Apoptosis assessed by flow cytometry analysis was used to evaluate the role of Ars2 in cells proliferation. The lentivirus-mediated gene knockdown approach was conducted to determine the function of Ars2. The orthotopic glioblastoma xenograft was used to demonstrate the role of Ars2 in glioblastoma growth in vivo. The high expression of Ars2 was observed in several glioblastoma cell lines and was significantly associated with poorer overall survival. Importantly, the overexpression of Ars2 promoted cell proliferation and colony formation in glioblastoma cells, whereas the depletion of Ars2 inhibited cell proliferation, colony formation, and tumor growth. Mechanistic study revealed that knockdown of Ars2 reduced the expression levels of miR-6798-3p, which was responsible for the up-regulation of p53 and p21, leading to apoptosis. Furthermore, the knockdown of Ars2 suppressed tumor growth in orthotopic glioblastoma xenograft model and significantly prolonged the survival time of the tumor-bearing mice. These findings identify a critical role for Ars2 in regulation of proliferation and tumorigenicity in glioblastoma and suggest that Ars2 could be a critical therapeutic target for glioblastoma intervention.

Flavonoids inhibit cell proliferation and induce apoptosis and autophagy through downregulation of PI3Kγ mediated PI3K/AKT/mTOR/p70S6K/ULK signaling pathway in human breast cancer cells.

Anticancer activities of flavonoids derived from Tephroseris kirilowii (Turcz.) Holub. were evaluated in human cancer cells. We isolated and identified, for the first time, eight flavonoids from T. kirilowii and found that three of them (IH: isorhamnetin, GN: genkwanin, and Aca: acacetin) inhibited cell proliferation in a variety of human cancer cell lines. These active flavonoids caused cell cycle arrest at G2/M phase and induced apoptosis and autophagy in human breast cancer cells. Molecular docking revealed that these flavonoids dock in the ATP binding pocket of PI3Kγ. Importantly, treatment with these flavonoids decreased the levels of PI3Kγ-p110, phospho-PI3K, phospho-AKT, phospho-mTOR, phospho-p70S6K, and phospho-ULK. Pretreatment with PI3Kγ specific inhibitor AS605240 potentiated flavonoids-mediated inactivation of AKT, mTOR, p70S6K, ULK, and apoptosis. Taken together, these findings represent a novel mechanism by which downregulation of PI3Kγ-p110 and consequent interruption of PI3K/AKT/mTOR/p70S6K/ULK signaling pathway might play a critical functional role in these flavonoids-induced cell cycle arrest at G2/M phase, apoptosis, and autophagy. Our studies provide novel insights into the anticancer activities of selected flavonoids and their potential uses in anticancer therapy.

The cyclohexene derivative MC-3129 exhibits antileukemic activity via RhoA/ROCK1/PTEN/PI3K/Akt pathway-mediated mitochondrial translocation of cofilin.

The effects of MC-3129, a synthetic cyclohexene derivative, on cell viability and apoptosis have been investigated in human leukemia cells. Exposure of leukemia cells to MC-3129 led to the inhibition of cell viability and induction of apoptosis through the dephosphorylation and mitochondrial translocation of cofilin. A mechanistic study revealed that interruption of the RhoA/ROCK1/PTEN/PI3K/Akt signaling pathway plays a crucial role in the MC-3129-mediated dephosphorylation and mitochondrial translocation of cofilin and induction of apoptosis. Our in vivo study also showed that the MC-3129-mediated inhibition of the tumor growth in a mouse leukemia xenograft model is associated with the interruption of ROCK1/PTEN/PI3K/Akt signaling and apoptosis. Molecular docking suggested that MC-3129 might activate the RhoA/ROCK1 pathway by targeting LPAR2. Collectively, these findings suggest a hierarchical model, in which the induction of apoptosis by MC-3129 primarily results from the activation of RhoA/ROCK1/PTEN and inactivation of PI3K/Akt, leading to the dephosphorylation and mitochondrial translocation of cofilin, and culminating in cytochrome c release, caspase activation, and apoptosis. Our study reveals a novel role for RhoA/ROCK1/PTEN/PI3K/Akt signaling in the regulation of mitochondrial translocation of cofilin and apoptosis and suggests MC-3129 as a potential drug for the treatment of human leukemia.

A novel autophagy inhibitor berbamine blocks SNARE-mediated autophagosome-lysosome fusion through upregulation of BNIP3.

Increasing evidences reveal that autophagy inhibitor could enhance the effect of chemotherapy to cancer. However, few autophagy inhibitors are currently approved for clinical application in humans. Berbamine (BBM) is a natural compound extracted from traditional Chinese medicine that is widely used for treatment of a variety of diseases without any obvious side effects. Here we found that BBM is a novel auophagy inhibitor, which potently induced the accumulation of autophagosomes by inhibiting autophagosome-lysosome fusion in human breast cancer cells. Mechanistically, we found that BBM blocked autophagosome-lysosome fusion by inhibiting the interaction of SNAP29 and VAMP8. Furthermore, BBM induced upregulation of BNIP3 and the interaction between SNAP29 and BNIP3. BNIP3 depletion or SNAP29 overexpression abrogated BBM-mediated blockade of autophagosome-lysosome fusion through the interaction between SNAP29 and VAMP8, whereas BNIP3 overexpression blocked autophagosome-lysosome fusion through inhibition of the interaction between SNAP29 and VAMP8. These findings suggest that upregulation of BNIP3 and interaction between BNIP3 and SNAP29 could be involved in BBM-mediated blockade of autophagosome-lysosome fusion through inhibition of the interaction between SNAP29 and VAMP8. Our findings identify the critical role of BNIP3 in blockade of autophagosome-lysosome fusion mediated by BBM, and suggest that BBM could potentially be further developed as a novel autophagy inhibitor, which could enhance the effect of chemotherapy to cancer.

Mitochondrial fission and mitophagy depend on cofilin-mediated actin depolymerization activity at the mitochondrial fission site.

Mitochondria fission and mitophagy are fundamentally crucial to cellular physiology and play important roles in cancer progression. Developing a comprehensive understanding of the molecular mechanism underlying mitochondrial fission and mitophagy will provide novel strategies for cancer prevention and treatment. Actin has been shown to participate in mitochondrial fission and mitophagy regulation. Cofilin is best known as an actin-depolymerizing factor. However, the molecular mechanism by which cofilin regulates mitochondrial fission and mitophagy remains largely unknown. Here we report that knockdown of cofilin attenuates and overexpression of cofilin potentiates mitochondrial fission as well as PINK1/PARK2-dependent mitophagy induced by staurosporine (STS), etoposide (ETO), and carbonyl cyanide 3-chlorophenylhydrazone (CCCP). Cofilin-mediated-PINK1 (PTEN-induced putative kinase 1) accumulation mainly depends on its regulation of mitochondrial proteases, including peptidase mitochondrial processing beta (MPPß), presenilin-associated rhomboid-like protease (PARL), and ATPase family gene 3-like 2 (AFG3L2), via mitochondrial membrane potential activity. We also found that the interaction and colocalization of G-actin/F-actin with cofilin at mitochondrial fission sites undergo constriction after CCCP treatment. Pretreatment with the actin polymerization inhibitor latrunculin B (LatB) increased and actin-depolymerization inhibitor jasplakinolide (Jas) decreased mitochondrial translocation of actin induced by STS, ETO, and CCCP. Both LatB and Jas abrogated CCCP-mediated mitochondrial fission and mitophagy. Our data suggest that G-actin is the actin form that is translocated to mitochondria, and the actin-depolymerization activity regulated by cofilin at the mitochondrial fission site is crucial for inducing mitochondrial fission and mitophagy.

Polyphyllin I induces mitophagic and apoptotic cell death in human breast cancer cells by increasing mitochondrial PINK1 levels.

The molecular mechanisms underlying the anti-breast cancer effects of polyphyllin I, a natural compound extracted from Paris polyphylla rhizomes, are not fully understood. In the present study, we found that polyphyllin I induces mitochondrial translocation of DRP1 by dephosphorylating DRP1 at Ser637, leading to mitochondrial fission, cytochrome c release from mitochondria into the cytosol and, ultimately apoptosis. Polyphyllin I also increased the stabilization of full-length PINK1 at the mitochondrial surface, leading to the recruitment of PARK2, P62, ubiquitin, and LC3B-II to mitochondria and culminating in mitophagy. PINK1 knockdown markedly suppressed polyphyllin I-induced mitophagy and enhanced polyphyllin I-induced, DRP1-dependent mitochondrial fission and apoptosis. Furthermore, suppression of DRP1 by mdivi-1 or shRNA inhibited PINK1 knockdown/polyphyllin I-induced mitochondrial fragmentation and apoptosis, suggesting that PINK1 depletion leads to excessive fission and, subsequently, mitochondrial fragmentation. An in vivo study confirmed that polyphyllin I greatly inhibited tumor growth and induced apoptosis in MDA-MB-231 xenografts, and these effects were enhanced by PINK1 knockdown. These data describe the mechanism by which PINK1 contributes to polyphyllin I-induced mitophagy and apoptosis and suggest that polyphyllin I may be an effective drug for breast cancer treatment.