Development of chimeric antigen receptor (CAR)-T cells targeting A56 viral protein implanted by oncolytic virus.

To address the challenge of solid tumor targeting in CAR-T therapy, we utilized the A56 antigen, which is uniquely expressed on a diverse range of cancer cells following the systemic administration of an oncolytic vaccinia virus (OVV). Immunohistochemical assays precisely confirmed exclusive localization of A56 to tumor tissues. In vitro studies demonstrated a distinct superiority of A56-dependent CAR-T cytotoxicity across multiple cancer cell lines. Building on these in vitro observations, we strategically administered A56 CAR-T cells, OVV, and hydroxyurea (HU) combination in HCT-116 tumor-bearing non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice, leading to a significant reduction in tumor size and an extended time to progression. Consequently, A56-targeting combinatorial immunotherapy provides the benefit of reducing inadvertent CAR-T effects on normal cells while preserving its effectiveness against cancer cells. Furthermore, our approach of implanting A56 via OVV on tumors facilitates a wide therapeutic application of CAR-T cells across various solid tumors.

MHY446 induces apoptosis via reactive oxygen species-mediated endoplasmic reticulum stress in HCT116 human colorectal cancer cells.

This study investigated the potential of a newly synthesized histone deacetylase (HDAC) inhibitor, MHY446, in inducing cell death in HCT116 colorectal cancer cells and compared its activity with that of suberoylanilide hydroxamic acid (SAHA), a well-known HDAC inhibitor. The results showed that MHY446 increased the acetylation of histones H3 and H4 and decreased the expression and activity of HDAC proteins in HCT116 cells. Additionally, MHY446 was confirmed to bind more strongly to HDAC1 than HDAC2 and inhibit its activity. In vivo experiments using nude mice revealed that MHY446 was as effective as SAHA in inhibiting HCT116 cell-grafted tumor growth. This study also evaluated the biological effects of MHY446 on cell survival and death pathways. The reactive oxygen species (ROS) scavenger N-acetyl-L-cysteine (NAC) confirmed that ROS play a role in MHY446-induced cell death by reducing poly(ADP-ribose) polymerase cleavage. MHY446 also induced cell death via endoplasmic reticulum (ER) stress by increasing the expression of ER stress-related proteins. NAC treatment decreased the expression of ER stress-related proteins, indicating that ROS mediate ER stress as an upstream signaling pathway and induce cell death. While MHY446 did not exhibit superior HDAC inhibition efficacy compared to SAHA, it is anticipated to provide innovative insights into the future development of therapeutic agents for human CRC by offering novel chemical structure-activity relationship-related information.

Pathogenesis, Intervention, and Current Status of Drug Development for Sarcopenia: A Review.

Sarcopenia refers to the loss of muscle strength and mass in older individuals and is a major determinant of fall risk and impaired ability to perform activities of daily living, often leading to disability, loss of independence, and death. Owing to its impact on morbidity, mortality, and healthcare expenditure, sarcopenia in the elderly has become a major focus of research and public policy debates worldwide. Despite its clinical importance, sarcopenia remains under-recognized and poorly managed in routine clinical practice, partly owing to the lack of available diagnostic testing and uniform diagnostic criteria. Since the World Health Organization and the United States assigned a disease code for sarcopenia in 2016, countries worldwide have assigned their own disease codes for sarcopenia. However, there are currently no approved pharmacological agents for the treatment of sarcopenia; therefore, interventions for sarcopenia primarily focus on physical therapy for muscle strengthening and gait training as well as adequate protein intake. In this review, we aimed to examine the latest information on the epidemiology, molecular mechanisms, interventions, and possible treatments with new drugs for sarcopenia.

Recent Developments in Combination Chemotherapy for Colorectal and Breast Cancers with Topoisomerase Inhibitors.

DNA topoisomerases are important enzymes that stabilize DNA supercoiling and resolve entanglements. There are two main types of topoisomerases in all cells: type I, which causes single-stranded DNA breaks, and type II, which cuts double-stranded DNA. Topoisomerase activity is particularly increased in rapidly dividing cells, such as cancer cells. Topoisomerase inhibitors have been an effective chemotherapeutic option for the treatment of several cancers. In addition, combination cancer therapy with topoisomerase inhibitors may increase therapeutic efficacy and decrease resistance or side effects. Topoisomerase inhibitors are currently being used worldwide, including in the United States, and clinical trials on the combination of topoisomerase inhibitors with other drugs are currently underway. The primary objective of this review was to comprehensively analyze the current clinical landscape concerning the combined application of irinotecan, an extensively investigated type I topoisomerase inhibitor for colorectal cancer, and doxorubicin, an extensively researched type II topoisomerase inhibitor for breast cancer, while presenting a novel approach for cancer therapy.

Therapeutic Potential of Bioactive Components from Scutellaria baicalensis Georgi in Inflammatory Bowel Disease and Colorectal Cancer: A Review.

Scutellaria baicalensis Georgi (SBG), an herbal medicine with various biological activities, including anti-inflammatory, anticancer, antiviral, antibacterial, and antioxidant activities, is effective in treatment of colitis, hepatitis, pneumonia, respiratory infections, and allergic diseases. This herbal medicine consists of major active substances, such as baicalin, baicalein, wogonoside, and wogonin. Inflammatory bowel disease (IBD) comprises a group of inflammatory conditions of the colon and small intestine, with Crohn's disease and ulcerative colitis being the main types. IBD can lead to serious complications, such as increased risk of colorectal cancer (CRC), one of the most common cancers worldwide. Currently, there is no cure for IBD, and its incidence has been increasing over the past few decades. This review comprehensively summarizes the efficacy of SBG in IBD and CRC and may serve as a reference for future research and development of drugs for IBD and cancer treatment.

MHY2251, a New SIRT1 Inhibitor, Induces Apoptosis via JNK/p53 Pathway in HCT116 Human Colorectal Cancer Cells.

Sirtuins (SIRTs) belong to the nicotinamide adenine dinucleotide (NAD+)-dependent class III histone deacetylase family. They are key regulators of cellular and physiological processes, such as cell survival, senescence, differentiation, DNA damage and stress response, cellular metabolism, and aging. SIRTs also influence carcinogenesis, making them potential targets for anticancer therapeutic strategies. In this study, we investigated the anticancer properties and underlying molecular mechanisms of a novel SIRT1 inhibitor, MHY2251, in human colorectal cancer (CRC) cells. MHY2251 reduced the viability of various human CRC cell lines, especially those with wild-type TP53. MHY2251 inhibited SIRT1 activity and SIRT1/2 protein expression, while promoting p53 acetylation, which is a target of SIRT1 in HCT116 cells. MHY2251 treatment triggered apoptosis in HCT116 cells. It increased the percentage of late apoptotic cells and the sub-G1 fraction (as detected by flow cytometric analysis) and induced DNA fragmentation. In addition, MHY2251 upregulated the expression of FasL and Fas, altered the ratio of Bax/Bcl-2, downregulated the levels of pro-caspase-8, -9, and -3 proteins, and induced subsequent poly(ADP-ribose) polymerase cleavage. The induction of apoptosis by MHY2251 was related to the activation of the caspase cascade, which was significantly attenuated by pre-treatment with Z-VAD-FMK, a pan-caspase inhibitor. Furthermore, MHY2251 stimulated the phosphorylation of c-Jun N-terminal kinase (JNK), and MHY2251-triggered apoptosis was blocked by pre-treatment with SP600125, a JNK inhibitor. This finding indicated the specific involvement of JNK in MHY2251-induced apoptosis. MHY2251 shows considerable potential as a therapeutic agent for targeting human CRC via the inhibition of SIRT1 and activation of JNK/p53 pathway.

Mechanism of Resveratrol-Induced Programmed Cell Death and New Drug Discovery against Cancer: A Review.

Resveratrol (3,5,4'-trihydroxy-trans-stilbene), a polyphenol found in grapes, red wine, peanuts, and apples, has been reported to exhibit a wide range of biological and pharmacological properties. In addition, resveratrol has been reported to intervene in multiple stages of carcinogenesis. It has also been known to kill several human cancer cells through programmed cell death (PCD) mechanisms such as apoptosis, autophagy, and necroptosis. However, resveratrol has limitations in its use as an anticancer agent because it is susceptible to photoisomerization owing to its unstable double bond, short half-life, and is rapidly metabolized and eliminated. Trans-(E)-resveratrol is nontoxic, and has several biological and pharmacological activities. However, little is known about the pharmacological properties of the photoisomerized cis-(Z)-resveratrol. Therefore, many studies on resveratrol derivatives and analogues that can overcome the shortcomings of resveratrol and increase its anticancer activity are underway. This review comprehensively summarizes the literature related to resveratrol-induced PCD, such as apoptosis, autophagy, necroptosis, and the development status of synthetic resveratrol derivatives and analogues as novel anticancer drugs.

Mechanism of Bile Acid-Induced Programmed Cell Death and Drug Discovery against Cancer: A Review.

Bile acids are major signaling molecules that play a significant role as emulsifiers in the digestion and absorption of dietary lipids. Bile acids are amphiphilic molecules produced by the reaction of enzymes with cholesterol as a substrate, and they are the primary metabolites of cholesterol in the body. Bile acids were initially considered as tumor promoters, but many studies have deemed them to be tumor suppressors. The tumor-suppressive effect of bile acids is associated with programmed cell death. Moreover, based on this fact, several synthetic bile acid derivatives have also been used to induce programmed cell death in several types of human cancers. This review comprehensively summarizes the literature related to bile acid-induced programmed cell death, such as apoptosis, autophagy, and necroptosis, and the status of drug development using synthetic bile acid derivatives against human cancers. We hope that this review will provide a reference for the future research and development of drugs against cancer.

Role of Induced Programmed Cell Death in the Chemopreventive Potential of Apigenin.

The flavonoid apigenin (4',5,7-trihydroxyflavone), which is one of the most widely distributed phytochemicals in the plant kingdom, is one of the most thoroughly investigated phenolic components. Previous studies have attributed the physiological effects of apigenin to its anti-allergic, antibacterial, antidiabetic, anti-inflammatory, antioxidant, antiviral, and blood-pressure-lowering properties, and its documented anticancer properties have been attributed to the induction of apoptosis and autophagy, the inhibition of inflammation, angiogenesis, and cell proliferation, and the regulation of cellular responses to oxidative stress and DNA damage. The most well-known mechanism for the compound's anticancer effects in human cancer cell lines is apoptosis, followed by autophagy, and studies have also reported that apigenin induces novel cell death mechanisms, such as necroptosis and ferroptosis. Therefore, the aim of this paper is to review the therapeutic potential of apigenin as a chemopreventive agent, as well as the roles of programmed cell death mechanisms in the compound's chemopreventive properties.

MHY2245, a Sirtuin Inhibitor, Induces Cell Cycle Arrest and Apoptosis in HCT116 Human Colorectal Cancer Cells.

Sirtuins (SIRTs), which are nicotinamide adenine dinucleotide-dependent class III histone deacetylases, regulate cell division, survival, and senescence. Although sirtinol, a synthetic SIRT inhibitor, is known to exhibit antitumor effects, its mechanism of action is not well understood. Therefore, we aimed to assess the anticancer effects and underlying mechanism of MHY2245, a derivative of sirtinol, in HCT116 human colorectal cancer cells in vitro. Treatment with MHY2245 decreased SIRT1 activity and caused DNA damage, leading to the upregulation of p53 acetylation, and increased levels of p53, phosphorylation of H2A histone family member X, ataxia telangiectasia and Rad3-related kinase, checkpoint kinase 1 (Chk1), and Chk2. The level of the breast cancer type 1 susceptibility protein was also found to decrease. MHY2245 induced G2/M phase cell cycle arrest via the downregulation of cyclin B1, cell division cycle protein 2 (Cdc2), and Cdc25c. Further, MHY2245 induced HCT116 cell death via apoptosis, which was accompanied by internucleosomal DNA fragmentation, decreased B-cell lymphoma 2 (Bcl-2) levels, increased Bcl-2-asscociated X protein levels, cleavage of poly(ADP-ribose) polymerase, and activation of caspases -3, -8, and -9. Overall, MHY2245 induces cell cycle arrest, triggers apoptosis through caspase activation, and exhibits DNA damage response-associated anticancer effects.

Novel SIRT Inhibitor, MHY2256, Induces Cell Cycle Arrest, Apoptosis, and Autophagic Cell Death in HCT116 Human Colorectal Cancer Cells.

We examined the anticancer effects of a novel sirtuin inhibitor, MHY2256, on HCT116 human colorectal cancer cells to investigate its underlying molecular mechanisms. MHY2256 significantly suppressed the activity of sirtuin 1 and expression levels of sirtuin 1/2 and stimulated acetylation of forkhead box O1, which is a target protein of sirtuin 1. Treatment with MHY2256 inhibited the growth of the HCT116 (TP53 wild-type), HT-29 (TP53 mutant), and DLD-1 (TP53 mutant) human colorectal cancer cell lines. In addition, MHY2256 induced G0/G1 phase arrest of the cell cycle progression, which was accompanied by the reduction of cyclin D1 and cyclin E and the decrease of cyclin-dependent kinase 2, cyclin-dependent kinase 4, cyclin-dependent kinase 6, phosphorylated retinoblastoma protein, and E2F transcription factor 1. Apoptosis induction was shown by DNA fragmentation and increase in late apoptosis, which were detected using flow cytometric analysis. MHY2256 downregulated expression levels of procaspase-8, -9, and -3 and led to subsequent poly(ADP-ribose) polymerase cleavage. MHY2256-induced apoptosis was involved in the activation of caspase-8, -9, and -3 and was prevented by pretreatment with Z-VAD-FMK, a pan-caspase inhibitor. Furthermore, the autophagic effects of MHY2256 were observed as cytoplasmic vacuolation, green fluorescent protein-light-chain 3 punctate dots, accumulation of acidic vesicular organelles, and upregulated expression level of light-chain 3-II. Taken together, these results suggest that MHY2256 could be a potential novel sirtuin inhibitor for the chemoprevention or treatment of colorectal cancer or both.

Characterization of Oncolytic Vaccinia Virus Harboring the Human IFNB1 and CES2 Transgenes.

PURPOSE: The purpose of this study was to assess characteristics of SJ-815, a novel oncolytic vaccinia virus lacking a functional thymidine kinase-encoding TK gene, and instead, having two human transgenes: the IFNB1 that encodes interferon ß1, and the CES2 that encodes carboxylesterase 2, which metabolizes the prodrug, irinotecan, into cytotoxic SN-38. MATERIALS AND METHODS: Viral replication and dissemination of SJ-815 were measured by plaque assay and comet assay, respectively, and compared to the backbone of SJ-815, a modified Western Reserve virus named WI. Tumor cytotoxicity of SJ-815 (or mSJ-815, which has the murine IFNB1 transgene for mouse cancers) was evaluated using human and mouse cancer cells. Antitumor effects of SJ-815, with/without irinotecan, were evaluated using a human pancreatic cancer-bearing mouse model and a syngeneic melanoma-bearing mouse model. The SN-38/ irinotecan ratios in mouse melanoma tissue 4 days post irinotecan treatment were compared between groups with and without SJ-815 intravenous injection. RESULTS: SJ-815 demonstrated significantly lower viral replication and dissemination, but considerably stronger in vitro tumor cytotoxicity than WI. The combination use of SJ-815 plus irinotecan generated substantial tumor regression in the human pancreatic cancer model, and significantly prolonged survival in the melanoma model (hazard ratio, 0.11; 95% confidence interval, 0.02 to 0.50; p=0.013). The tumor SN-38/irinotecan ratios were over 3-fold higher in the group with SJ-815 than those without (p < 0.001). CONCLUSION: SJ-815 demonstrates distinct characteristics gained from the inserted IFNB1 and CES2 transgenes. The potent antitumor effects of SJ-815, particularly when combined with irinotecan, against multiple solid tumors make SJ-815 an attractive candidate for further preclinical and clinical studies.

Pro-apoptotic effect of the novel benzylidene derivative MHY695 in human colon cancer cells.

The induction of apoptosis is a useful strategy in anti-cancer research. Various Moon Hyung Yang (MHY) compounds have been developed as novel anti-cancer drug candidates; in the present study, the pro-apoptotic effects of (Z)-5-(3-ethoxy-4- hydroxybenzylidene)-2-thioxothiazolidin-4-one (MHY695) on HCT116 human colon cancer cells were assessed. MTT assays were performed to investigate the dose-dependent cytotoxic effects of MHY695 on HCT116 cells. Immunofluorescence staining and flow cytometry analyses were performed to identify apoptotic cell death, and western blot analysis was used to investigate the apoptotic-signaling pathways. A mouse xenograft model was also used to determine the effects of MHY695 in vivo. MHY695 decreased the viability of HCT116 cells and induced apoptotic cytotoxicity. The apoptotic mechanisms induced by MHY695 involved the dephosphorylation of Bcl-2-associated agonist of cell death protein following protein kinase B inactivation, induced myeloid leukaemia cell differentiation protein and BH3-interacting domain death agonist truncation, caspase-3 and -9 activation and poly (ADP-ribose) polymerase cleavage. In addition, MHY695 significantly suppressed tumor growth in the mouse xenograft model, compared with the vehicle control. Notably, MHY695 exhibited potent anti-cancer effects in four different types of human colon cancer cell line, including Caco-2, DLD-1, HT-29 and HCT116. Additionally, MHY695 showed reduced cytotoxicity in NCM460, normal colonic epithelial cells. Furthermore, MHY-induced cytotoxicity in colon cancer cells was independent of the tumor suppressor protein p53. Collectively, these observations suggested that MHY695 may be a novel drug for the treatment of colon cancer.

PKM2 Knockdown Induces Autophagic Cell Death via AKT/mTOR Pathway in Human Prostate Cancer Cells.

BACKGROUND/AIMS: Pyruvate kinase M2 (PKM2) is essential for aerobic glycolysis. Although high PKM2 expression is observed in various cancer tissues, its functional role in cancer metabolism is unclear. Here, we investigated the role of PKM2 in regulating autophagy and its associated pathways in prostate cancer cells. METHODS: Immunohistochemistry was performed to compare the expression level of PKM2 in prostate cancer patients and normal human, whereas expression of PKM2 in several cell lines was also examined by using western blot. PKM2 expression was silenced using various small interfering RNAs (siRNAs). Cell viability was examined using IncuCyte ZOOM™ live cell imaging system. Western blotting and immunofluorescence were performed to investigate the PKM2 knockdown on other cellular signaling molecules. Acridine orange and Monodansylcadaverine staining was performed to check effect of PKM2 knockdown on autophagy induction. High performance thin layer chromatography was carried out to quantify the level of different cellular metabolites (pyruvate and lactate). Colony formation assay was performed to determine the ability of a cells to form large colonies. RESULTS: PKM2 was highly expressed in prostate cancer patients as compared to normal human. PKM2 siRNA-transfected prostate cancer cells showed significantly reduced viability. Acridine orange, Monodansylcadaverine staining and western blotting analysis showed that PKM2 downregulation markedly increased autophagic cell death. Results of western blotting analysis showed that PKM2 knockdown affected protein kinase B/mechanistic target of rapamycin 1 pathway, which consequently downregulated the expression of glycolytic enzymes lactate dehydrogenase A and glucose transporter 1. Knockdown of PKM2 also reduced the colony formation ability of human prostate cancer cell DU145. CONCLUSION: To the best of our knowledge, this is the first study to show that PKM2 inhibition alters prostate cancer cell metabolism and induces autophagy, thus providing new perspectives for developing PKM2-targeting anticancer therapies for treating prostate cancer.

Redefining Chronic Inflammation in Aging and Age-Related Diseases: Proposal of the Senoinflammation Concept.

Age-associated chronic inflammation is characterized by unresolved and uncontrolled inflammation with multivariable low-grade, chronic and systemic responses that exacerbate the aging process and age-related chronic diseases. Currently, there are two major hypotheses related to the involvement of chronic inflammation in the aging process: molecular inflammation of aging and inflammaging. However, neither of these hypotheses satisfactorily addresses age-related chronic inflammation, considering the recent advances that have been made in inflammation research. A more comprehensive view of age-related inflammation, that has a scope beyond the conventional view, is therefore required. In this review, we discuss newly emerging data on multi-phase inflammatory networks and proinflammatory pathways as they relate to aging. We describe the age-related upregulation of nuclear factor (NF)-κB signaling, cytokines/chemokines, endoplasmic reticulum (ER) stress, inflammasome, and lipid accumulation. The later sections of this review present our expanded view of age-related senescent inflammation, a process we term "senoinflammation", that we propose here as a novel concept. As described in the discussion, senoinflammation provides a schema highlighting the important and ever-increasing roles of proinflammatory senescence-associated secretome, inflammasome, ER stress, TLRs, and microRNAs, which support the senoinflammation concept. It is hoped that this new concept of senoinflammation opens wider and deeper avenues for basic inflammation research and provides new insights into the anti-inflammatory therapeutic strategies targeting the multiple proinflammatory pathways and mediators and mediators that underlie the pathophysiological aging process.

Morin enhances auranofin anticancer activity by up-regulation of DR4 and DR5 and modulation of Bcl-2 through reactive oxygen species generation in Hep3B human hepatocellular carcinoma cells.

Evidence suggests that auranofin (AF) exhibits anticancer activity by inhibiting thioredoxin reductase (TrxR). Here, in this study, we have investigated the synergistic effects of AF and morin and their mechanism for the anticancer effects focusing on apoptosis in Hep3B human hepatocellular carcinoma cells. We assessed the anticancer activities by annexin V/PI double staining, caspase, and TrxR activity assay. Morin enhances the inhibitory effects on TrxR activity of AF as well as reducing cell viability. Annexin V/PI double staining revealed that morin/AF cotreatment induced apoptotic cell death. Morin enhances AF-induced mitochondrial membrane potential (ΔΨm) loss and cytochrome c release. Further, morin/AF cotreatment upregulated death receptor DR4/DR5, modulated Bcl-2 family members (upregulation of Bax and downregulation of Bcl-2), and activated caspase-3, -8, and -9. Morin also enhances AF-induced reactive oxygen species (ROS) generation. The anticancer effects results from caspase-dependent apoptosis, which was triggered via extrinsic pathway by upregulating TRAIL receptors (DR4/DR5) and enhanced via intrinsic pathway by modulating Bcl-2 and inhibitor of apoptosis protein family members. These are related to ROS generation. In conclusion, this study provides evidence that morin can enhance the anticancer activity of AF in Hep3B human hepatocellular carcinoma cells, indicating that its combination could be an alternative treatment strategy for the hepatocellular carcinoma.

Roles of folate in skeletal muscle cell development and functions.

Folate is the generic term for both naturally occurring food folate and folic acid, the fully oxidized monoglutamate form of the vitamin that is used in dietary supplements and fortified foods. It is a water-soluble vitamin B9 and is important for health, growth, and development. As a precursor of various cofactors, folate is required for one-carbon donors in the synthesis of DNA bases and other essential biomolecules. A lack of dietary folate can lead to folate deficiency and can therefore result in several health problems, including macrocytic anemia, elevated plasma homocysteine, cardiovascular disease, birth defects, carcinogenesis, muscle weakness, and difficulty in walking. Several studies have implied that folate might exert a positive effect on skeletal muscle development. However, the precise effects of folate in skeletal muscle development are still poorly understood. Thus, this review provides an updated discussion of the roles of folate in skeletal muscle cell development and the effects of folic acid supplementation on the functions of skeletal muscle cells.

Preclinical safety evaluation of hepatic arterial infusion of oncolytic poxvirus.

PURPOSE: Oncolytic poxvirus has shown promise in treating various solid tumors, such as liver cancer, and administration of oncolytic poxvirus via the hepatic artery may provide more survival benefits than other routes of administration. However, there is a lack of safety information to guide the application of hepatic arterial infusion (HAI) of oncolytic poxvirus in human studies. To investigate the acute and chronic toxicity of HAI administration of oncolytic poxvirus in animals and provide safety information for future human studies. METHODS: VVtk-, a vaccinia poxvirus with inactivated thymidine kinase gene, was administered via HAI to rabbits with normal liver function under angiography (1×108 or 1×109 pfu), and rats with N-nitrosomorpholine-induced precancerous liver cirrhosis under open surgery (1×108 pfu). Body weights and survival were monitored and blood samples were collected for hematological and biochemical tests. Distribution of A56 (a specific marker for poxvirus infection) in rabbit organs was evaluated using immunofluorescence assays. RESULTS: HAI of high doses of VVtk- did not cause any acute or chronic changes in body weight, survival or in biochemical, hematological tests in the 2 animal models, and none of the changes showed dose dependency (in rabbit study), or were influenced by liver cirrhosis (in rat study). A56 was not detected in any of the major rabbit organs. CONCLUSION: HAI may provide a safe alternative route of oncolytic poxvirus administration for human studies.

MHY440, a Novel Topoisomerase Ι Inhibitor, Induces Cell Cycle Arrest and Apoptosis via a ROS-Dependent DNA Damage Signaling Pathway in AGS Human Gastric Cancer Cells.

We investigated the antitumor activity and action mechanism of MHY440 in AGS human gastric cancer cells. MHY440 inhibited topoisomerase (Topo) Ι activity and was associated with a DNA damage response signaling pathway. It exhibited a stronger anti-proliferative effect on AGS cells relative to Hs27 human foreskin fibroblast cells, and this effect was both time- and concentration-dependent. MHY440 also increased cell arrest in the G2/M phase by decreasing cyclin B1, Cdc2, and Cdc25c, and upregulating p53 and p73. MHY440 induced AGS cell apoptosis through the upregulation of Fas-L, Fas, and Bax as well as the proteolysis of BH3 interacting-domain death agonist and poly(ADP-ribose) polymerase. It also contributed to the loss of mitochondrial membrane potential. The apoptotic cell death induced by MHY440 was inhibited by pretreatment with Z-VAD-FMK, a pan-caspase inhibitor, indicating that apoptosis was caspase-dependent. Moreover, the apoptotic effect of MHY440 was reactive oxygen species (ROS)-dependent, as evidenced by the inhibition of MHY440-induced PARP cleavage and ROS generation via N-acetylcysteine-induced ROS scavenging. Taken together, MHY440 showed anticancer effects by inhibiting Topo I, regulating the cell cycle, inducing apoptosis through caspase activation, and generating ROS, suggesting that MHY440 has considerable potential as a therapeutic agent for human gastric cancer.

Induction of apoptosis by ethanol extract of Citrus unshiu Markovich peel in human bladder cancer T24 cells through ROS-mediated inactivation of the PI3K/Akt pathway.

Citrus unshiu peel has been used to prevent and treat various diseases in traditional East-Asian medicine including in Korea. Extracts of C. unshiu peel are known to have various pharmacological effects including antioxidant, anti-inflammatory, and antibacterial properties. Although the possibility of their anti-cancer activity has recently been reported, the exact mechanisms in human cancer cells have not been sufficiently studied. In this study, the inhibitory effect of ethanol extract of C. unshiu peel (EECU) on the growth of human bladder cancer T24 cells was evaluated and the underlying mechanism was investigated. The present study demonstrated that the suppression of T24 cell viability by EECU is associated with apoptosis induction. EECU-induced apoptosis was found to correlate with an activation of caspase-8, -9, and -3 in concomitance with a decrease in the expression of the inhibitor of apoptosis family of proteins and an increase in the Bax:Bcl-2 ratio accompanied by the proteolytic degradation of poly(ADP-ribose) polymerase. EECU also increased the generation of reactive oxygen species (ROS), collapse of mitochondrial membrane potential, and cytochrome c release to the cytosol, along with a truncation of Bid. In addition, EECU inactivated phosphatidylinositol 3-kinase (PI3K) as well as Akt, a downstream molecular target of PI3K, and LY294002, a specific PI3K inhibitor significantly enhanced EECU-induced apoptosis and cell viability reduction. However, N-acetyl cysteine, a general ROS scavenger, completely reversed the EECU-induced dephosphorylation of PI3K and Akt, as well as cell apoptosis. Taken together, these findings suggest that EECU inhibits T24 cell proliferation by activating intrinsic and extrinsic apoptosis pathways through a ROS-mediated inactivation of the PI3K/Akt pathway.