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Several marine drugs exert anticancer effects by inducing oxidative stress, which becomes overloaded and kills cancer cells when redox homeostasis is imbalanced. The downregulation of antioxidant signaling induces oxidative stress, while its upregulation attenuates oxidative stress. Marine drugs have miRNA-modulating effects against cancer cells. However, the potential antioxidant targets of such drugs have been rarely explored. This review aims to categorize the marine-drug-modulated miRNAs that downregulate their antioxidant targets, causing oxidative stress in anticancer treatments. We also categorize the downregulation of oxidative-stress-inducing miRNAs in antioxidant protection among non-cancer cells. We summarize the putative antioxidant targets of miRNA-modulating marine drugs by introducing a bioinformatics tool (miRDB). Finally, the marine drugs affecting antioxidant targets are surveyed. In this way, the connections between marine drugs and their modulating miRNA and antioxidant targets are innovatively categorized to provide a precise network for exploring their potential anticancer functions and protective effects on non-cancer cells., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Protein phosphatase 2A (PP2A), a heterotrimeric holoenzyme (scaffolding, catalytic, and regulatory subunits), regulates dephosphorylation for more than half of serine/threonine phosphosites and exhibits diverse cellular functions. Although several studies on natural products and miRNAs have emphasized their impacts on PP2A regulation, their connections lack systemic organization. Moreover, only part of the PP2A family has been investigated. This review focuses on the PP2A-modulating effects of natural products and miRNAs' interactions with potential PP2A targets in cancer and non-cancer cells. PP2A-modulating natural products and miRNAs were retrieved through a literature search. Utilizing the miRDB database, potential PP2A targets of these PP2A-modulating miRNAs for the whole set (17 members) of the PP2A family were retrieved. Finally, PP2A-modulating natural products and miRNAs were linked via a literature search. This review provides systemic directions for assessing natural products and miRNAs relating to the PP2A-modulating functions in cancer and disease treatments., (© 2024 Wiley Periodicals LLC.)

Ferroptosis, which comprises iron-dependent cell death, is crucial in cancer and non-cancer treatments. Exosomes, the extracellular vesicles, may deliver biomolecules to regulate disease progression. The interplay between ferroptosis and exosomes may modulate cancer development but is rarely investigated in natural product treatments and their modulating miRNAs. This review focuses on the ferroptosis-modulating effects of natural products and miRNAs concerning their participation in ferroptosis and exosome biogenesis (secretion and assembly)-related targets in cancer and non-cancer cells. Natural products and miRNAs with ferroptosis-modulating effects were retrieved and organized. Next, a literature search established the connection of a panel of ferroptosis-modulating genes to these ferroptosis-associated natural products. Moreover, ferroptosis-associated miRNAs were inputted into the miRNA database (miRDB) to bioinformatically search the potential targets for the modulation of ferroptosis and exosome biogenesis. Finally, the literature search provided a connection between ferroptosis-modulating miRNAs and natural products. Consequently, the connections from ferroptosis-miRNA-exosome biogenesis to natural product-based anticancer treatments are well-organized. This review sheds light on the research directions for integrating miRNAs and exosome biogenesis into the ferroptosis-modulating therapeutic effects of natural products on cancer and non-cancer diseases.

Antioral cancer drugs need a greater antiproliferative impact on cancer than on normal cells. Demethoxymurrapanine (DEMU) inhibits proliferation in several cancer cells, but an in-depth investigation was necessary. This study evaluated the proliferation-modulating effects of DEMU, focusing on oral cancer and normal cells. DEMU (0, 2, 3, and 4 μg/mL) at 48 h treatments inhibited the proliferation of oral cancer cells (the cell viability (%) for Ca9-22 cells was 100.0 ± 2.2, 75.4 ± 5.6, 26.0 ± 3.8, and 15.4 ± 1.4, and for CAL 27 cells was 100.0 ± 9.4, 77.2 ± 5.9, 57.4 ± 10.7, and 27.1 ± 1.1) more strongly than that of normal cells (the cell viability (%) for S-G cells was 100.0 ± 6.6, 91.0 ± 4.6, 95.0 ± 2.6, and 95.8 ± 5.5), although this was blocked by the antioxidant N-acetylcysteine. The presence of oxidative stress was evidenced by the increase of reactive oxygen species and mitochondrial superoxide and the downregulation of the cellular antioxidant glutathione in oral cancer cells, but these changes were minor in normal cells. DEMU also caused greater induction of the subG1 phase, extrinsic and intrinsic apoptosis (annexin V and caspases 3, 8, and 9), and DNA damage (γH2AX and 8-hydroxy-2-deoxyguanosine) in oral cancer than in normal cells. N-acetylcysteine attenuated all these DEMU-induced changes. Together, these data demonstrate the preferential antiproliferative function of DEMU in oral cancer cells, with the preferential induction of oxidative stress, apoptosis, and DNA damage in these cancer cells, and low cytotoxicity toward normal cells., (© 2023 Wiley Periodicals LLC.)

This in vitro study examines the anti-oral cancer effects and mechanisms of a combined X-ray/SK2 treatment, i.e., X-ray and 6- n -butoxy-10-nitro-12,13-dioxa-11-azatricyclo[7.3.1.0 2,7 ]trideca-2,4,6,10-tetraene (SK2). ATP cell viability and flow cytometry-based cell cycle, apoptosis, oxidative stress, and DNA damage assessments were conducted. The X-ray/SK2 treatment exhibited lower viability in oral cancer (Ca9-22 and CAL 27) cells than in normal (Smulow-Glickman, S-G) cells, i.e., 32.0%, 46.1% vs. 59.0%, which showed more antiproliferative changes than with X-ray or SK2 treatment. Oral cancer cells under X-ray/SK2 treatment showed slight subG1 and G2/M increments and induced high annexin V-monitored apoptosis compared to X-ray or SK2 treatment. The X-ray/SK2 treatment showed higher caspase 3 and 8 levels for oral cancer cells than other treatments. X-ray/SK2 showed a higher caspase 9 level in CAL 27 cells than other treatments, while Ca9-22 cells showed similar levels under X-ray and/or SK2. The X-ray/SK2 treatment showed higher reactive oxygen species (ROS) generation and mitochondrial membrane potential (MMP) depletion than other treatments. Meanwhile, the mitochondrial superoxide (MitoSOX) and glutathione levels in X-ray/SK2 treatment did not exhibit the highest rank compared to others. Moreover, oral cancer cells had higher γH2AX and/or 8-hydroxy-2-deoxyguanosine levels from X-ray/SK2 treatment than others. All these measurements for X-ray/SK2 in oral cancer cells were higher than in normal cells and attenuated by N-acetylcysteine. In conclusion, X-ray/SK2 treatment showed ROS-dependent enhanced antiproliferative, apoptotic, and DNA damage effects in oral cancer cells with a lower cytotoxic influence on normal cells.

Increased neddylation benefits the survival of several types of cancer cells. The inhibition of neddylation has the potential to exert anticancer effects but is rarely assessed in oral cancer cells. This study aimed to investigate the antiproliferation potential of a neddylation inhibitor MLN4924 (pevonedistat) for oral cancer cells. MLN4924 inhibited the cell viability of oral cancer cells more than that of normal oral cells (HGF-1) with 100% viability, that is, IC 50 values of oral cancer cells (CAL 27, OC-2, and Ca9-22) are 1.8, 1.4, and 1.9 μM. MLN4924 caused apoptotic changes such as the subG1 accumulation, activation of annexin V, pancaspase, and caspases 3/8/9 of oral cancer cells at a greater rate than in normal oral cells. MLN4924 induced greater oxidative stress in oral cancer cells compared to normal cells by upregulating reactive oxygen species and mitochondrial superoxide and depleting the mitochondrial membrane potential and glutathione. In oral cancer cells, preferential inductions also occurred for DNA damage (γH2AX and 8-oxo-2'-deoxyguanosine). Therefore, this investigation demonstrates that MLN4924 is a potential anti-oral-cancer agent showing preferential inhibition of apoptosis and promotion of DNA damage with fewer cytotoxic effects on normal cells., (© 2023 Wiley Periodicals LLC.)

Exosomes are cell-derived membranous structures primarily involved in the delivery of the payload to the recipient cells, and they play central roles in carcinogenesis and metastasis. Radiotherapy is a common cancer treatment that occasionally generates exosomal miRNA-associated modulation to regulate the therapeutic anticancer function and side effects. Combining radiotherapy and natural products may modulate the radioprotective and radiosensitizing responses of non-cancer and cancer cells, but there is a knowledge gap regarding the connection of this combined treatment with exosomal miRNAs and their downstream targets for radiation and exosome biogenesis. This review focuses on radioprotective natural products in terms of their impacts on exosomal miRNAs to target radiation-modulating and exosome biogenesis (secretion and assembly) genes. Several natural products have individually demonstrated radioprotective and miRNA-modulating effects. However, the impact of natural-product-modulated miRNAs on radiation response and exosome biogenesis remains unclear. In this review, by searching through PubMed/Google Scholar, available reports on potential functions that show radioprotection for non-cancer tissues and radiosensitization for cancer among these natural-product-modulated miRNAs were assessed. Next, by accessing the miRNA database (miRDB), the predicted targets of the radiation- and exosome biogenesis-modulating genes from the Gene Ontology database (MGI) were retrieved bioinformatically based on these miRNAs. Moreover, the target-centric analysis showed that several natural products share the same miRNAs and targets to regulate radiation response and exosome biogenesis. As a result, the miRNA-radiomodulation (radioprotection and radiosensitization)-exosome biogenesis axis in regard to natural-product-mediated radiotherapeutic effects is well organized. This review focuses on natural products and their regulating effects on miRNAs to assess the potential impacts of radiomodulation and exosome biogenesis for both the radiosensitization of cancer cells and the radioprotection of non-cancer cells.

Physalis plants are commonly used traditional medicinal herbs, and most of their extracts containing withanolides show anticancer effects. Physapruin A (PHA), a withanolide isolated from P. peruviana , shows antiproliferative effects on breast cancer cells involving oxidative stress, apoptosis, and autophagy. However, the other oxidative stress-associated response, such as endoplasmic reticulum (ER) stress, and its participation in regulating apoptosis in PHA-treated breast cancer cells remain unclear. This study aims to explore the function of oxidative stress and ER stress in modulating the proliferation and apoptosis of breast cancer cells treated with PHA. PHA induced a more significant ER expansion and aggresome formation of breast cancer cells (MCF7 and MDA-MB-231). The mRNA and protein levels of ER stress-responsive genes ( IRE1α and BIP ) were upregulated by PHA in breast cancer cells. The co-treatment of PHA with the ER stress-inducer (thapsigargin, TG), i.e., TG/PHA, demonstrated synergistic antiproliferation, reactive oxygen species generation, subG1 accumulation, and apoptosis (annexin V and caspases 3/8 activation) as examined by ATP assay, flow cytometry, and western blotting. These ER stress responses, their associated antiproliferation, and apoptosis changes were partly alleviated by the N -acetylcysteine, an oxidative stress inhibitor. Taken together, PHA exhibits ER stress-inducing function to promote antiproliferation and apoptosis of breast cancer cells involving oxidative stress.

Triple-negative breast cancer (TNBC) is insensitive to target therapy for non-TNBC and needs novel drug discovery. Extracts of the traditional herb Boesenbergia plant in Southern Asia exhibit anticancer effects and contain novel bioactive compounds but merely show cytotoxicity. We recently isolated a new compound from B. stenophylla , stenophyllol B (StenB), but the impact and mechanism of its proliferation-modulating function on TNBC cells remain uninvestigated. This study aimed to assess the antiproliferative responses of StenB in TNBC cells and examine the drug safety in normal cells. StenB effectively suppressed the proliferation of TNBC cells rather than normal cells in terms of an ATP assay. This preferential antiproliferative function was alleviated by pretreating inhibitors for oxidative stress ( N -acetylcysteine (NAC)) and apoptosis (Z-VAD-FMK). Accordingly, the oxidative-stress-related mechanisms were further assessed. StenB caused subG1 and G2/M accumulation but reduced the G1 phase in TNBC cells, while normal cells remained unchanged between the control and StenB treatments. The apoptosis behavior of TNBC cells was suppressed by StenB, whereas that of normal cells was not suppressed according to an annexin V assay. StenB-modulated apoptosis signaling, such as for caspases 3, 8, and 9, was more significantly activated in TNBC than in normal cells. StenB also caused oxidative stress in TNBC cells but not in normal cells according to a flow cytometry assay monitoring reactive oxygen species, mitochondrial superoxide, and their membrane potential. StenB induced greater DNA damage responses (γH2AX and 8-hydroxy-2-deoxyguanosine) in TNBC than in normal cells. All these StenB responses were alleviated by NAC pretreatment. Collectively, StenB modulated oxidative stress responses, leading to the antiproliferation of TNBC cells with little cytotoxicity in normal cells.

Cancerous exosomes contain diverse biomolecules that regulate cancer progression. Modulating exosome biogenesis with clinical drugs has become an effective strategy for cancer therapy. Suppressing exosomal processing (assembly and secretion) may block exosomal function to reduce the proliferation of cancer cells. However, the information on natural products that modulate cancer exosomes lacks systemic organization, particularly for exosomal long noncoding RNAs (lncRNAs). There is a gap in the connection between exosomal lncRNAs and exosomal processing. This review introduces the database (LncTarD) to explore the potential of exosomal lncRNAs and their sponging miRNAs. The names of sponging miRNAs were transferred to the database (miRDB) for the target prediction of exosomal processing genes. Moreover, the impacts of lncRNAs, sponging miRNAs, and exosomal processing on the tumor microenvironment (TME) and natural-product-modulating anticancer effects were then retrieved and organized. This review sheds light on the functions of exosomal lncRNAs, sponging miRNAs, and exosomal processing in anticancer processes. It also provides future directions for the application of natural products when regulating cancerous exosomal lncRNAs.

Ginger-derived compounds are abundant sources of anticancer natural products. However, the anticancer effects of ( E )-3-hydroxy-1-(4'-hydroxy-3',5'-dimethoxyphenyl)-tetradecan-6-en-5-one (3HDT) have not been examined. This study aims to assess the antiproliferation ability of 3HDT on triple-negative breast cancer (TNBC) cells. 3HDT showed dose-responsive antiproliferation for TNBC cells (HCC1937 and Hs578T). Moreover, 3HDT exerted higher antiproliferation and apoptosis on TNBC cells than on normal cells (H184B5F5/M10). By examining reactive oxygen species, mitochondrial membrane potential, and glutathione, we found that 3HDT provided higher inductions for oxidative stress in TNBC cells compared with normal cells. Antiproliferation, oxidative stress, antioxidant signaling, and apoptosis were recovered by N -acetylcysteine, indicating that 3HDT preferentially induced oxidative-stress-mediated antiproliferation in TNBC cells but not in normal cells. Moreover, by examining γH2A histone family member X (γH2AX) and 8-hydroxy-2-deoxyguanosine, we found that 3HDT provided higher inductions for DNA damage, which was also reverted by N -acetylcysteine. In conclusion, 3HDT is an effective anticancer drug with preferential antiproliferation, oxidative stress, apoptosis, and DNA damage effects on TNBC cells.

Manoalide provides preferential antiproliferation of oral cancer but is non-cytotoxic to normal cells by modulating reactive oxygen species (ROS) and apoptosis. Although ROS interplays with endoplasmic reticulum (ER) stress and apoptosis, the influence of ER stress on manoalide-triggered apoptosis has not been reported. The role of ER stress in manoalide-induced preferential antiproliferation and apoptosis was assessed in this study. Manoalide induces a higher ER expansion and aggresome accumulation of oral cancer than normal cells. Generally, manoalide differentially influences higher mRNA and protein expressions of ER-stress-associated genes ( PERK , IRE1α , ATF6 , and BIP ) in oral cancer cells than in normal cells. Subsequently, the contribution of ER stress on manoalide-treated oral cancer cells was further examined. ER stress inducer, thapsigargin, enhances the manoalide-induced antiproliferation, caspase 3/7 activation, and autophagy of oral cancer cells rather than normal cells. Moreover, N -acetylcysteine, an ROS inhibitor, reverses the responses of ER stress, aggresome formation, and the antiproliferation of oral cancer cells. Consequently, the preferential ER stress of manoalide-treated oral cancer cells is crucial for its antiproliferative effect.

Many miRNAs are known to target the AKT serine-threonine kinase (AKT) pathway, which is critical for the regulation of several cell functions in cancer cell development. Many natural products exhibiting anticancer effects have been reported, but their connections to the AKT pathway (AKT and its effectors) and miRNAs have rarely been investigated. This review aimed to demarcate the relationship between miRNAs and the AKT pathway during the regulation of cancer cell functions by natural products. Identifying the connections between miRNAs and the AKT pathway and between miRNAs and natural products made it possible to establish an miRNA/AKT/natural product axis to facilitate a better understanding of their anticancer mechanisms. Moreover, the miRNA database (miRDB) was used to retrieve more AKT pathway-related target candidates for miRNAs. By evaluating the reported facts, the cell functions of these database-generated candidates were connected to natural products. Therefore, this review provides a comprehensive overview of the natural product/miRNA/AKT pathway in the modulation of cancer cell development.

Cancer-derived exosomes exhibit sophisticated functions, such as proliferation, apoptosis, migration, resistance, and tumor microenvironment changes. Several clinical drugs modulate these exosome functions, but the impacts of natural products are not well understood. Exosome functions are regulated by exosome processing, such as secretion and assembly. The modulation of these exosome-processing genes can exert the anticancer and precancer effects of cancer-derived exosomes. This review focuses on the cancer-derived exosomal miRNAs that regulate exosome processing, acting on the natural-product-modulating cell functions of cancer cells. However, the role of exosomal processing has been overlooked in several studies of exosomal miRNAs and natural products. In this study, utilizing the bioinformatics database (miRDB), the exosome-processing genes of natural-product-modulated exosomal miRNAs were predicted. Consequently, several natural drugs that modulate exosome processing and exosomal miRNAs and regulate cancer cell functions are described here. This review sheds light on and improves our understanding of the modulating effects of exosomal miRNAs and their potential exosomal processing targets on anticancer treatments based on the use of natural products.

The anticancer effects and mechanisms of marine sponge Aaptos suberitoides were rarely assessed, especially for methanol extract of A. suberitoides (MEAS) to breast cancer cells. This study evaluated the differential suppression effects of proliferation by MEAS between breast cancer and normal cells. MEAS demonstrated more antiproliferation impact on breast cancer cells than normal cells, indicating oxidative stress-dependent preferential antiproliferation effects on breast cancer cells but not for normal cells. Several oxidative stress-associated responses were highly induced by MEAS in breast cancer cells but not normal cells, including the generations of cellular and mitochondrial oxidative stress as well as the depletion of mitochondrial membrane potential. MEAS downregulated cellular antioxidants such as glutathione, partly contributing to the upregulation of oxidative stress in breast cancer cells. This preferential oxidative stress generation is accompanied by more DNA damage (γH2AX and 8-hydroxy-2-deoxyguanosine) in breast cancer cells than in normal cells. N -acetylcysteine reverted these MEAS-triggered responses. In conclusion, MEAS is a potential natural product for treating breast cancer cells with the characteristics of preferential antiproliferation function without cytotoxicity to normal cells in vitro.

Background: Keloid is a benign tumor with high recurrence rate; accordingly, complete surgical excision with adjuvant radiotherapy is one of the most effective treatments. This study reviewed outcomes of keloid patients receiving surgery and adjuvant radiotherapy in Kaohsiung Medical University Hospital., Materials and Methods: All patients received radiation dose with 15 Gy, with their first radiotherapy within 24 hours after surgical excision. The end points were recurrence rate and local recurrence-free interval (LRFI), defined clinically as palpable gross tumor over the treatment site and duration from the last day of radiotherapy to disease recurrence., Results: From May 2017 to July 2020, 32 patients with 40 keloid lesions were included. The mean age for these patients was 37.6 years, and the median follow-up time was 15.3 months. The overall recurrence rate was 52.5%, and the median LRFI was 9.7 months. Recurrence rates for males and females were 46.7% and 56% ( P = 0.567), respectively; for head and ear, chest, shoulder and upper extremities, and abdomen and back were 12.5%, 61.5%, 63.6%, and 62.5% ( P = 0.093); for lesions over 20 cm 2 and below 20 cm 2 were 62.5% and 50% ( P = 0.527); and for megavoltage electron beam and kilovoltage photon beam were 56.7% and 40% ( P = 0.361), respectively. Patients were further classified into 2 groups by lesion sites, which showed lower recurrence rate ( P = 0.011) and longer LRFI ( P = 0.028) with lesions over the head and ear than other sites., Conclusions: We found that lesion site might be a prognostic factor for keloid recurrence. Adjuvant radiation dose escalation for high-recurrence risk areas (other than the head and ear) might be required., Competing Interests: Conflicts of interest and sources of funding: none declared., (Copyright © 2022 The Author(s). Published by Wolters Kluwer Health, Inc.)

Physapruin A (PHA), a Physalis peruviana -derived withanolide, exhibits antiproliferation activity against oral and breast cancer cells. However, its potential antitumor effects in combined treatments remain unclear. This investigation focused on evaluating the impact of the combined treatment of ultraviolet-C with PHA (UVC/PHA) on the proliferation of oral cancer cells. The UVC-caused antiproliferation was enhanced by combination with PHA in oral cancer (Ca9-22 and CAL 27) but not normal cells (SG), as evidenced by ATP detection, compared with UVC or PHA alone. UVC/PHA showed a greater extent of subG1 increase, G2/M arrest, annexin-V-assessed apoptosis, caspase 3/7 activation, and reactive oxygen species (ROS) in the UVC or PHA treatment of oral cancer compared to normal cells. Moreover, the mitochondrial functions, such as mitochondrial superoxide bursts and mitochondrial membrane potential destruction, of oral cancer cells were also enhanced by UVC/PHA compared to UVC or PHA alone. These oxidative stresses triggered γH2AX and 8-hydroxyl-2'-deoxyguanosine-assessed DNA damage to a greater extent under UVC/PHA treatment than under UVC or PHA treatment alone. The ROS inhibitor N -acetylcysteine reversed all these UVC/PHA-promoted changes. In conclusion, UVC/PHA is a promising strategy for decreasing the proliferation of oral cancer cells but shows no inhibitory effect on normal cells.

Background/purpose: High-dose-rate (HDR) electronic brachytherapy (EBT) has been shown to be effective for non-melanoma skin cancer (NMSC) in Caucasian patients. However, its efficacy remains unknown in Asian patients. To analyze the clinical outcome of HDR EBT for NMSC in a Taiwanese medical center., Methods: Medical records over a 5-year period between January 2015 to December 2019 were retrospectively analyzed., Results: Forty-seven patients with 54 NMSC including 42 basal cell carcinomas (BCCs) and 12 squamous cell carcinomas (SCCs) were treated with HDR EBT. The average age was 73.8 years. The mean radiation dose was 45.3 Gy (40-80 Gy). Mean follow-up duration was 33.1 months. Adequate local control was achieved in 50 lesions (92.6%). Grade 1 acute skin toxicity was noted in 63.0% of lesions, while no tumors had Grade 4 acute toxicity. No ulceration was observed six months after completion of treatment. At the last follow-up visit, all lesions were rated to have "fair" to "excellent" cosmetic outcomes., Conclusion: HDR EBT provides adequate clinical outcomes and cosmetic results for NMSC in Asian patients. Further investigation of the dosage guidelines is needed for Asian patients with NMSC., Competing Interests: Declaration of competing interest The authors have no conflicts of interest relevant to this article., (Copyright © 2022. Published by Elsevier B.V.)

A novel nitrated [6,6,6]tricycles-derived compound containing nitro, methoxy, and ispropyloxy groups, namely SK1, was developed in our previous report. However, the anticancer effects of SK1 were not assessed. Moreover, SK1 contains two nitro groups (NO 2 ) and one nitrogen-oxygen (N-O) bond exhibiting the potential for oxidative stress generation, but this was not examined. The present study aimed to evaluate the antiproliferation effects and oxidative stress and its associated responses between oral cancer and normal cells. Based on the MTS assay, SK1 demonstrated more antiproliferation ability in oral cancer cells than normal cells, reversed by N -acetylcysteine. This suggests that SK1 causes antiproliferation effects preferentially in an oxidative stress-dependent manner. The oxidative stress-associated responses were further validated, showing higher ROS/MitoSOX burst, MMP, and GSH depletion in oral cancer cells than in normal cells. Meanwhile, SK1 caused oxidative stress-causing apoptosis, such as caspases 3/8/9, and DNA damages, such as γH2AX and 8-OHdG, to a greater extent in oral cancer cells than in normal cells. Siilar to cell viability, these oxidative stress responses were partially diminished by NAC, indicating that SK1 promoted oxidative stress-dependent responses. In conclusion, SK1 exerts oxidative stress, apoptosis, and DNA damage to a greater extent to oral cancer cells than in normal cells.

The purpose of this study aimed to assess the antiproliferation effects of methanol extract of T. swinhoei (METS) and explore the detailed responses of oral cancer cells compared to normal cells. METS effectively inhibits the cell proliferation of oral cancer cells but does not affect normal cell viability, exhibiting preferential antiproliferation function. METS exerted more subG1 accumulation, apoptosis induction, cellular and mitochondrial oxidative stress, and DNA damage than normal cells, reverted by oxidative stress inhibitor N -acetylcysteine. This METS-caused oxidative stress was validated to attribute to the downregulation of glutathione. METS activated both extrinsic and intrinsic caspases. DNA double-strand breaks (γH2AX) and oxidative DNA damage (8-hydroxy-2-deoxyguanosine) were stimulated by METS. Therefore, for the first time, this investigation shed light on exploring the functions and responses of preferential antiproliferation of METS in oral cancer cells.

AKT serine-threonine kinase (AKT) and its effectors are essential for maintaining cell proliferation, apoptosis, autophagy, endoplasmic reticulum (ER) stress, mitochondrial morphogenesis (fission/fusion), ferroptosis, necroptosis, DNA damage response (damage and repair), senescence, and migration of cancer cells. Several lncRNAs and circRNAs also regulate the expression of these functions by numerous pathways. However, the impact on cell functions by lncRNAs and circRNAs regulating AKT and its effectors is poorly understood. This review provides comprehensive information about the relationship of lncRNAs and circRNAs with AKT on the cell functions of cancer cells. the roles of several lncRNAs and circRNAs acting on AKT effectors, such as FOXO, mTORC1/2, S6K1/2, 4EBP1, SREBP, and HIF are explored. To further validate the relationship between AKT, AKT effectors, lncRNAs, and circRNAs, more predicted AKT- and AKT effector-targeting lncRNAs and circRNAs were retrieved from the LncTarD and circBase databases. Consistently, using an in-depth literature survey, these AKT- and AKT effector-targeting database lncRNAs and circRNAs were related to cell functions. Therefore, some lncRNAs and circRNAs can regulate several cell functions through modulating AKT and AKT effectors. This review provides insights into a comprehensive network of AKT and AKT effectors connecting to lncRNAs and circRNAs in the regulation of cancer cell functions.

Oxidative stress and AKT serine-threonine kinase (AKT) are responsible for regulating several cell functions of cancer cells. Several natural products modulate both oxidative stress and AKT for anticancer effects. However, the impact of natural product-modulating oxidative stress and AKT on cell functions lacks systemic understanding. Notably, the contribution of regulating cell functions by AKT downstream effectors is not yet well integrated. This review explores the role of oxidative stress and AKT pathway (AKT/AKT effectors) on ten cell functions, including apoptosis, autophagy, endoplasmic reticulum stress, mitochondrial morphogenesis, ferroptosis, necroptosis, DNA damage response, senescence, migration, and cell-cycle progression. The impact of oxidative stress and AKT are connected to these cell functions through cell function mediators. Moreover, the AKT effectors related to cell functions are integrated. Based on this rationale, natural products with the modulating abilities for oxidative stress and AKT pathway exhibit the potential to regulate these cell functions, but some were rarely reported, particularly for AKT effectors. This review sheds light on understanding the roles of oxidative stress and AKT pathway in regulating cell functions, providing future directions for natural products in cancer treatment.

Data regarding the effects of crude extract of Commelina plants in oral cancer treatment are scarce. This present study aimed to assess the proliferation-modulating effects of the Commelina sp. (MECO) methanol extract on oral cancer cells in culture, Ca9-22, and CAL 27. MECO suppressed viability to a greater extent in oral cancer cells than in normal cells. MECO also induced more annexin V, apoptosis, and caspase signaling for caspases 3/8/9 in oral cancer cells. The preferential antiproliferation and apoptosis were associated with cellular and mitochondrial oxidative stress in oral cancer cells. Moreover, MECO also preferentially induced DNA damage in oral cancer cells by elevating γH2AX and 8-hydroxyl-2'-deoxyguanosine. The oxidative stress scavengers N -acetylcysteine or MitoTEMPO reverted these preferential antiproliferation mechanisms. It can be concluded that MECO is a natural product with preferential antiproliferation effects and exhibits an oxidative stress-associated mechanism in oral cancer cells.

Combined treatment is a promising anticancer strategy for improving antiproliferation compared with a single treatment but is limited by adverse side effects on normal cells. Fucoidan (FN), a brown-algae-derived polysaccharide safe food ingredient, exhibits preferential function for antiproliferation to oral cancer but not normal cells. Utilizing the preferential antiproliferation, the impacts of FN in regulating ultraviolet C (UVC) irradiation were assessed in oral cancer cells. A combined treatment (UVC/FN) reduced cell viability of oral cancer cells (Ca9-22 and CAL 27) more than single treatments (FN or UVC), i.e., 53.7%/54.6% vs. 71.2%/91.6%, and 89.2%/79.4%, respectively, while the cell viability of UVC/FN treating on non-malignant oral (S-G) was higher than oral cancer cells, ranging from 106.0 to 108.5%. Mechanistically, UVC/FN preferentially generated higher subG1 accumulation and apoptosis-related inductions (annexin V, caspases 3, 8, and 9) in oral cancer cells than single treatments. UVC/FN preferentially generated higher oxidative stress than single treatments, as evidenced by flow cytometry-detecting reactive oxygen species, mitochondrial superoxide, and glutathione. Moreover, UVC/FN preferentially caused more DNA damage (γH2AX and 8-hydroxy-2'-deoxyguanosine) in oral cancer cells than in single treatments. N -acetylcysteine pretreatment validated the oxidative stress effects in these UVC/FN-induced changes. Taken together, FN effectively enhances UVC-triggered antiproliferation to oral cancer cells. UVC/FN provides a promising potential for preferential and synergistic antiproliferation in antioral cancer therapy.

The selective antiproliferation to oral cancer cells of Physalis peruviana -derived physapruin A (PHA) is rarely reported. Either drug-induced apoptosis and DNA damage or DNA repair suppression may effectively inhibit cancer cell proliferation. This study examined the selective antiproliferation ability of PHA and explored detailed mechanisms of apoptosis, DNA damage, and repair. During an ATP assay, PHA provided high cytotoxicity to two oral cancer cell lines (CAL 27 and Ca9-22) but no cytotoxicity to two non-malignant oral cells (HGF-1 and SG). This selective antiproliferation of PHA was associated with the selective generation of reactive oxygen species (ROS) in oral cancer cells rather than in non-malignant oral cells, as detected by flow cytometry. Moreover, PHA induced other oxidative stresses in oral cancer cells, such as mitochondrial superoxide generation and mitochondrial membrane potential depletion. PHA also demonstrated selective apoptosis in oral cancer cells rather than non-malignant cells in annexin V/7-aminoactinmycin D and caspase 3/7 activity assays. In flow cytometry and immunofluorescence assays, PHA induced γH2AX expressions and increased the γH2AX foci number of DNA damages in oral cancer cells. In contrast, the mRNA expressions for DNA repair signaling, including homologous recombination (HR) and non-homologous end joining (NHEJ)-associated genes, were inhibited by PHA in oral cancer cells. Moreover, the PHA-induced changes were alleviated by the oxidative stress inhibitor N -acetylcysteine. Therefore, PHA generates selective antiproliferation, oxidative stress, and apoptosis associated with DNA damage induction and DNA repair suppression in oral cancer cells.

Noncoding RNAs (ncRNAs) regulation of various diseases including cancer has been extensively studied. Reactive oxidative species (ROS) elevated by oxidative stress are associated with cancer progression and drug resistance, while autophagy serves as an ROS scavenger in cancer cells. However, the regulatory effects of ncRNAs on autophagy and ROS in various cancer cells remains complex. Here, we explore how currently investigated ncRNAs, mainly miRNAs and lncRNAs, are involved in ROS production through modulating antioxidant genes. The regulatory effects of miRNAs and lncRNAs on autophagy-related (ATG) proteins to control autophagy activity in cancer cells are discussed. Moreover, differential expression of ncRNAs in tumor and normal tissues of cancer patients are further analyzed using The Cancer Genome Atlas (TCGA) database. This review hypothesizes links between ATG genes- or antioxidant genes-modulated ncRNAs and ROS production, which might result in tumorigenesis, malignancy, and cancer recurrence. A better understanding of the regulation of ROS and autophagy by ncRNAs might advance the use of ncRNAs as diagnostic and prognostic markers as well as therapeutic targets in cancer therapy., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Physalis peruviana -derived physapruin A (PHA) is a potent compound that selectively generates reactive oxygen species (ROS) and induces cancer cell death. Autophagy, a cellular self-clearance pathway, can be induced by ROS and plays a dual role in cancer cell death. However, the role of autophagy in PHA-treated cancer cells is not understood. Our study initially showed that autophagy inhibitors such as bafilomycin A1 enhanced the cytotoxic effects of PHA in breast cancer cell lines, including MCF7 and MDA-MB-231. PHA treatment decreased the p62 protein level and increased LC3-II flux. PHA increased the fluorescence intensity of DAPGreen and DALGreen, which are used to reflect the formation of autophagosome/autolysosome and autolysosome, respectively. ROS scavenger N -acetylcysteine (NAC) decreased PHA-elevated autophagy activity, implying that PHA-induced ROS may be required for autophagy induction in breast cancer cells. Moreover, the autophagy inhibitor increased ROS levels and enhanced PHA-elevated ROS levels, while NAC scavenges the produced ROS resulting from PHA and autophagy inhibitor. In addition, the autophagy inhibitor elevated the PHA-induced proportion of annexin V/7-aminoactinmycin D and cleavage of caspase-3/8/9 and poly (ADP-ribose) polymerase. In contrast, NAC and apoptosis inhibitor Z-VAD-FMK blocked the proportion of annexin V/7-aminoactinmycin D and the activation of caspases. Taken together, PHA induced ROS to promote autophagy, which might play an antioxidant and anti-apoptotic role in breast cancer cells.

Combined treatment is an effective strategy to improve anticancer therapy, but severe side effects frequently limit this application. Drugs inhibiting the proliferation of cancer cells, but not normal cells, display preferential antiproliferation to cancer cells. It shows the benefits of avoiding side effects and enhancing antiproliferation for combined treatment. Nitrated [6,6,6]tricycles derivative (SK2), a novel chemical exhibiting benzo-fused dioxabicyclo[3.3.1]nonane core with an n -butyloxy substituent, exhibiting preferential antiproliferation, was chosen to evaluate its potential antioral cancer effect in vitro by combining it with ultraviolet C (UVC) irradiation. Combination treatment (UVC/SK2) caused lower viability in oral cancer cells (Ca9-22 and OC-2) than single treatment (20 J/m 2 UVC or 10 μg/mL SK2), i.e., 42.3%/41.1% vs. 81.6%/69.2%, and 89.5%/79.6%, respectively. In contrast, it showed a minor effect on cell viability of normal oral cells (HGF-1), ranging from 82.2 to 90.6%. Moreover, UVC/SK2 caused higher oxidative stress in oral cancer cells than normal cells through the examination of reactive oxygen species, mitochondrial superoxide, and mitochondrial membrane potential. UVC/SK2 also caused subG1 increment associated with apoptosis detections by assessing annexin V; panaspase; and caspases 3, 8, and 9. The antiproliferation and oxidative stress were reverted by N -acetylcysteine, validating the involvement of oxidative stress in antioral cancer cells. UVC/SK2 also caused DNA damage by detecting γH2AX and 8-hydroxy-2'-deoxyguanosine in oral cancer cells. In conclusion, SK2 is an effective enhancer for improving the UVC-caused antiproliferation against oral cancer cells in vitro. UVC/SK2 demonstrated a preferential and synergistic antiproliferation ability towards oral cancer cells with little adverse effects on normal cells.

SK2, a nitrated [6,6,6]tricycle derivative with an n -butyloxy group, showed selective antiproliferation effects on oral cancer but not on normal oral cells. This investigation assessed for the first time the synergistic antiproliferation potential of cisplatin/SK2 in oral cancer cells. Cell viability assay at 24 h showed that a low dose of combined cisplatin/SK2 (10 μM/10 μg/mL) provided more antiproliferation than cisplatin or SK2 alone. Cisplatin/SK2 triggered also more apoptosis inductions in terms of subG1 accumulation, annexin V, pancaspase, and caspase 3/8/9 measurements. Moreover, cisplatin/SK2 provided more oxidative stress and DNA damage in oral cancer cells than independent treatments. Oxidative stress inhibitors rescued the cisplatin/SK2-induced antiproliferation and oxidative stress generation. Moreover, cisplatin/SK2 induced more antiproliferation, apoptosis, oxidative stress, and DNA damage in oral cancer cells than in normal oral cells (S-G). In conclusion, low-dose cisplatin/SK2 combined treatment promoted selective and synergistic antiproliferation in oral cancer cells depending on oxidative-stress-associated responses.

The critical factors for regulating cancer metabolism are oxidative stress and phosphoinositide-3-kinase/AKT serine-threonine kinase/mechanistic target of the rapamycin kinase (PI3K/AKT/mTOR). However, the metabolic impacts of oxidative stress and PI3K/AKT/mTOR on individual mechanisms such as glycolysis (Warburg effect), pentose phosphate pathway (PPP), fatty acid synthesis, tricarboxylic acid cycle (TCA) cycle, glutaminolysis, and oxidative phosphorylation (OXPHOS) are complicated. Therefore, this review summarizes the individual and interacting functions of oxidative stress and PI3K/AKT/mTOR on metabolism. Moreover, natural products providing oxidative stress and PI3K/AKT/mTOR modulating effects have anticancer potential. Using the example of brown algae-derived fucoidan, the roles of oxidative stress and PI3K/AKT/mTOR were summarized, although their potential functions within diverse metabolisms were rarely investigated. We propose a potential application that fucoidan may regulate oxidative stress and PI3K/AKT/mTOR signaling to modulate their associated metabolic regulations. This review sheds light on understanding the impacts of oxidative stress and PI3K/AKT/mTOR on metabolism and the future direction of metabolism-based cancer therapy of fucoidan.

Fucoidan is a dietary brown algae-derived fucose-rich polysaccharide. However, the anticancer effects of fucoidan for oral cancer treatment remain unclear, particularly in terms of its preferential antiproliferation ability and oxidative-stress-associated responses. This study first evaluated the effects and mechanisms of the preferential antiproliferation of fucoidan between oral cancer and non-malignant oral cells (S-G). In a 48 h MTS assay, fucoidan showed higher antiproliferation in response to five types of oral cancer cells, but not S-G cells, demonstrating preferential antiproliferation of oral cancer cells. Oral cancer cells (Ca9-22 and CAL 27) showing high sensitivity to fucoidan were selected to explore the antiproliferation mechanism compared to S-G cells. Fucoidan showed subG1 accumulation and an annexin V increase in apoptosis, accompanied by caspase 8, 9, and 3 activations in oral cancer cells, but not in S-G cells. Fucoidan increased reactive oxygen species and mitochondrial superoxide levels and decreased cellular glutathione in oral cancer cells compared with S-G cells. These oxidative stress effects were attributed to the downregulation of antioxidant signaling genes ( NRF2 , TXN , and HMOX1 ) in oral cancer cells rather than S-G cells. Fucoidan showed DNA damage-inducible effects (γH2AX and 8-hydroxy-2-deoxyguanosine) in oral cancer cells but not in S-G cells. Accordingly, these preferential changes in oral cancer but not in non-malignant cells contribute to the preferential antiproliferation mechanism of fucoidan. Furthermore, these changes were reverted by pretreatment with the antioxidant N -acetylcysteine. Therefore, for the first time, this study provides a detailed understanding of the preferential antiproliferation effects and mechanisms of fucoidan in oral cancer cells.

Ginger is a popular spice and consists of several bioactive antioxidant compounds. Gingerenone A (Gin A), a novel compound isolated from Zingiber officinale , is rarely investigated for its anti-breast-cancer properties. Some ginger extracts have been reported to initiate senescence, an anticancer strategy. However, the anticancer effects of Gin A on breast cancer cells remain unclear. The present study aims to assess the modulating impact of Gin A acting on proliferation and senescence to breast cancer cells. Gin A diminished the cellular ATP content and decreased the cell viability of the MTS assay in several breast cancer cell lines. It also showed a delayed G2/M response to breast cancer cells (MCF7 and MDA-MB-231). N -acetylcysteine (NAC), an oxidative stress inhibitor, can revert these responses of antiproliferation and G2/M delay. The oxidative stress and senescence responses of Gin A were further validated by increasing reactive oxygen species, mitochondrial superoxide, and β-galactosidase activity, which were reverted by NAC. Gin A also upregulated senescence-associated gene expressions. In addition to oxidative stress, Gin A also induced DNA damage responses by increasing γH2AX level and foci and generating 8-hydroxyl-2'-deoxyguanosine in breast cancer cells, which were reverted by NAC. Therefore, Gin A promotes antiproliferation and senescence of breast cancer cells induced by oxidative stress.

Oxidative stress and the AKT serine/threonine kinase (AKT) signaling pathway are essential regulators in cellular migration, metastasis, and angiogenesis. More than 300 withanolides were discovered from the plant family Solanaceae, exhibiting diverse functions. Notably, the relationship between oxidative stress, AKT signaling, and angiogenesis in withanolide treatments lacks comprehensive understanding. Here, we summarize connecting evidence related to oxidative stress, AKT signaling, and angiogenesis in the zebrafish model. A convenient vertebrate model monitored the in vivo effects of developmental and tumor xenograft angiogenesis using zebrafish embryos. The oxidative stress and AKT-signaling-modulating abilities of withanolides were highlighted in cancer treatments, which indicated that further assessments of their angiogenesis-modulating potential are necessary in the future. Moreover, targeting AKT for inhibiting AKT and its AKT signaling shows the potential for anti-migration and anti-angiogenesis purposes for future application to withanolides. This particularly holds for investigating the anti-angiogenetic effects mediated by the oxidative stress and AKT signaling pathways in withanolide-based cancer therapy in the future.

Cryptocaryone (CPC) was previously reported as preferential for killing natural products in oral cancer cells. However, its radiosensitizing potential combined with ultraviolet C (UVC) cell killing of oral cancer cells remains unclear. This study evaluates the combined anti-proliferation effect and clarifies the mechanism of combined UVC/CPC effects on oral cancer cells. UVC/CPC shows higher anti-proliferation than individual and control treatments in a low cytotoxic environment on normal oral cells. Mechanistically, combined UVC/CPC generates high levels of reactive oxygen species and induces mitochondrial dysfunction by generating mitochondrial superoxide, increasing mitochondrial mass and causing the potential destruction of the mitochondrial membrane compared to individual treatments. Moreover, combined UVC/CPC causes higher G2/M arrest and triggers apoptosis, with greater evidence of cell cycle disturbance, annexin V, pancaspase, caspases 3/7 expression or activity in oral cancer cells than individual treatments. Western blotting further indicates that UVC/CPC induces overexpression for cleaved types of poly (ADP-ribose) polymerase and caspase 3 more than individual treatments. Additionally, UVC/CPC highly induces γH2AX and 8-hydroxy-2'-deoxyguanosine adducts as DNA damage in oral cancer cells. Taken together, CPC shows a radiosensitizing anti-proliferation effect on UVC irradiated oral cancer cells with combined effects through oxidative stress, apoptosis and DNA damage.

The benzo-fused dioxabicyclo[3.3.1]nonane core is the central framework in several natural products. Using this core, we had developed a novel nitrated [6,6,6]tricycle-derived compound containing an n -butyloxy group, namely, SK2. The anticancer potential of SK2 was not assessed. This study aimed to determine the antiproliferative function and investigated possible mechanisms of SK2 acting on oral cancer cells. SK2 preferentially killed oral cancer cells but caused no harmful effect on non-malignant oral cells. After the SK2 exposure of oral cancer cells, cells in the sub-G1 phase accumulated. This apoptosis-like outcome of SK2 treatment was validated to be apoptosis via observing an increasing annexin V population. Mechanistically, apoptosis signalers such as pancaspase, caspases 8, caspase 9, and caspase 3 were activated by SK2 in oral cancer cells. SK2 induced oxidative-stress-associated changes. Furthermore, SK2 caused DNA damage (γH2AX and 8-hydroxy-2'-deoxyguanosine). In conclusion, a novel nitrated [6,6,6]tricycle-derived compound, SK2, exhibits a preferential antiproliferative effect on oral cancer cells, accompanied by apoptosis, oxidative stress, and DNA damage.

Dihydrosinularin (DHS) is an analog of soft coral-derived sinularin; however, the anticancer effects and mechanisms of DHS have seldom been reported. This investigation examined the antiproliferation ability and mechanisms of DHS on oral cancer cells. In a cell viability assay, DHS showed growth inhibition against several types of oral cancer cell lines (Ca9-22, SCC-9, OECM-1, CAL 27, OC-2, and HSC-3) with no cytotoxic side effects on non-malignant oral cells (HGF-1). Ca9-22 and SCC-9 cell lines showing high susceptibility to DHS were selected to explore the antiproliferation mechanisms of DHS. DHS also causes apoptosis as detected by annexin V, pancaspase, and caspase 3 activation. DHS induces oxidative stress, leading to the generation of reactive oxygen species (ROS)/mitochondrial superoxide (MitoSOX) and mitochondrial membrane potential (MitoMP) depletion. DHS also induced DNA damage by probing γH2AX phosphorylation. Pretreatment with the ROS scavenger N -acetylcysteine (NAC) can partly counter these DHS-induced changes. We report that the marine natural product DHS can inhibit the cell growth of oral cancer cells. Exploring the mechanisms of this cancer cell growth inhibition, we demonstrate the prominent role DHS plays in oxidative stress.

Ethyl acetate Nepenthes extract (EANT) from Nepenthes thorellii × (ventricosa × maxima) shows antiproliferation and apoptosis but not necrosis in breast cancer cells, but this has not been investigated in oral cancer cells. In the present study, EANT shows no cytotoxicity to normal oral cells but exhibits selective killing to six oral cancer cell lines. They were suppressed by pretreatment of the antioxidant inhibitor N -acetylcysteine (NAC), demonstrating that EANT-induced cell death was mediated by oxidative stress. Concerning high sensitivity to EANT, Ca9-22 and CAL 27 oral cancer cells were chosen for exploring detailed selective killing mechanisms. EANT triggers a mixture of necrosis and apoptosis as determined by annexin V/7-aminoactinmycin D analysis. Still, they show differential switches from necrosis at a low (10 μg/mL) concentration to apoptosis at high (25 μg/mL) concentration of EANT in oral cancer cells. NAC induces necrosis but suppresses annexin V-detected apoptosis in oral cancer cells. Necrostatin 1 (NEC1), a necroptosis inhibitor, moderately suppresses necrosis but induces apoptosis at 10 μg/mL EANT. In contrast, Z-VAD-FMK, a pancaspase inhibitor, slightly causes necrosis but suppresses apoptosis at 10 μg/mL EANT. Furthermore, the flow cytometry-detected pancaspase activity is dose-responsively increased but is suppressed by NAC and ZVAD, although not for NEC1 in oral cancer cells. EANT causes several oxidative stress events such as reactive oxygen species, mitochondrial superoxide, and mitochondrial membrane depolarization. In response to oxidative stresses, the mRNA for antioxidant signaling, such as nuclear factor erythroid 2-like 2 ( NFE2L2 ), catalase ( CAT ), heme oxygenase 1 ( HMOX1 ), and thioredoxin ( TXN ), are overexpressed in oral cancer cells. Moreover, EANT also triggers DNA damage, as detected by γH2AX and 8-oxo-2'-deoxyguanosine adducts. The dependence of oxidative stress is validated by the evidence that NAC pretreatment reverts the changes of cellular and mitochondrial stress and DNA damage. Therefore, EANT exhibits antiproliferation involving an oxidative stress-dependent necrosis/apoptosis switch and DNA damage in oral cancer cells.

The anticancer effect of pomegranate polyphenolic extract POMx in oral cancer cells has rarely been explored, especially where its impact on mitochondrial functioning is concerned. Here, we attempt to evaluate the proliferation modulating function and mechanism of POMx against human oral cancer (Ca9-22, HSC-3, and OC-2) cells. POMx induced ATP depletion, subG1 accumulation, and annexin V/Western blotting-detected apoptosis in these three oral cancer cell lines but showed no toxicity to normal oral cell lines (HGF-1). POMx triggered mitochondrial membrane potential (MitoMP) disruption and mitochondrial superoxide (MitoSOX) generation associated with the differential downregulation of several antioxidant gene mRNA/protein expressions in oral cancer cells. POMx downregulated mitochondrial mass, mitochondrial DNA copy number, and mitochondrial biogenesis gene mRNA/protein expression in oral cancer cells. Moreover, POMx induced both PCR-based mitochondrial DNA damage and γH2AX-detected nuclear DNA damage in oral cancer cells. In conclusion, POMx provides antiproliferation and apoptosis of oral cancer cells through mechanisms of mitochondrial impairment.

Marine natural products are abundant resources for antioxidants, but the antioxidant property of the soft corals-derived sinularin and dihydrosinularin were unknown. This study aimed to assess antioxidant potential and antiproliferation effects of above compounds on cancer cells, and to investigate the possible relationships between them. Results show that sinularin and dihydrosinularin promptly reacted with 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2-azinobis (3-ethyl-benzothiazoline-6-sulfonic acid) (ABTS), and hydroxyl ( • OH), demonstrating a general radical scavenger activity. Sinularin and dihydrosinularin also show an induction for Fe +3 -reduction and Fe +2 -chelating capacity which both strengthen their antioxidant activities. Importantly, sinularin shows higher antioxidant properties than dihydrosinularin. Moreover, 24 h ATP assays show that sinularin leads to higher antiproliferation of breast, lung, and liver cancer cells than dihydrosinularin. Therefore, the differential antioxidant properties of sinularin and dihydrosinularin may contribute to their differential anti-proliferation of different cancer cells.

Combined treatment is increasingly used to improve cancer therapy. Non-ionizing radiation ultraviolet-C (UVC) and sinularin, a coral Sinularia flexibilis -derived cembranolide, were separately reported to provide an antiproliferation function to some kinds of cancer cells. However, an antiproliferation function using the combined treatment of UVC/sinularin has not been investigated as yet. This study aimed to examine the combined antiproliferation function and explore the combination of UVC/sinularin in oral cancer cells compared to normal oral cells. Regarding cell viability, UVC/sinularin displays the synergistic and selective killing of two oral cancer cell lines, but remains non-effective for normal oral cell lines compared to treatments in terms of MTS and ATP assays. In tests using the flow cytometry, luminescence, and Western blotting methods, UVC/sinularin-treated oral cancer cells exhibited higher reactive oxygen species production, mitochondrial superoxide generation, mitochondrial membrane potential destruction, annexin V, pan-caspase, caspase 3/7, and cleaved-poly (ADP-ribose) polymerase expressions than that in normal oral cells. Accordingly, oxidative stress and apoptosis are highly induced in a combined UVC/sinularin treatment. Moreover, UVC/sinularin treatment provides higher G2/M arrest and γH2AX/8-hydroxyl-2'deoxyguanosine-detected DNA damages in oral cancer cells than in the separate treatments. A pretreatment can revert all of these changes of UVC/sinularin treatment with the antioxidant N -acetylcysteine. Taken together, UVC/sinularin acting upon oral cancer cells exhibits a synergistic and selective antiproliferation ability involving oxidative stress-dependent apoptosis and cellular DNA damage with low toxic side effects on normal oral cells.

Breast cancer expresses clinically heterogeneous characteristics and requires multipurpose drug development for curing the different tumor subtypes. Many withanolides have been isolated from Physalis species showing anticancer effects, but the anticancer function of physapruin A (PHA) has rarely been investigated. In this study, the anticancer properties of PHA in breast cancer cells were examined by concentration and time-course experiments. In terms of cellular ATP content, PHA inhibited the proliferation of three kinds of breast cancer cells: MCF7 (estrogen receptor (ER)+, progesterone receptor (PR)+/-, human epidermal growth factor receptor 2 (HER2)-), SKBR3 (ER-/PR-/HER2+), and MDA-MB-231 (triple-negative). Moreover, PHA induced G2/M arrest in MCF7 and MDA-MB-231 cells. In terms of flow cytometry, PHA induced the generation of reactive oxygen species (ROS), the generation of mitochondrial superoxide, mitochondrial membrane potential depletion, and γH2AX-detected DNA damage in breast cancer MCF7 and MDA-MB-231 cells, which were suppressed by the ROS inhibitor N -acetylcysteine (NAC). In terms of flow cytometry and Western blotting, PHA induced apoptotic expression (annexin V, and intrinsic and extrinsic apoptotic signaling), which was suppressed by NAC and an apoptosis inhibitor (Z-VAD-FMK), in breast cancer cells. Therefore, PHA is a potential anti-breast-cancer natural product that modulates the oxidative-stress response, cell-cycle disturbance, apoptosis, and γH2AX-detected DNA damage.

We previously found that marine sponge-derived manoalide induced antiproliferation and apoptosis of oral cancer cells as well as reactive species generations probed by dichloro-dihydrofluorescein diacetate (DCFH-DA) and MitoSOX Red. However, the sources of cellular and mitochondrial redox stresses and the mutual interacting effects between these redox stresses and apoptosis remain unclear. To address this issue, we examined a panel of reactive species and used the inhibitors of cellular reactive species ( N -acetylcysteine (NAC)), mitochondrial reactive species (MitoTEMPO), and apoptosis (Z-VAD-FMK; ZVAD) to explore their interactions in manoalide-treated oral cancer Ca9-22 and CAL 27 cells. Hydroxyl (˙OH), nitrogen dioxide (NO 2 ˙), nitric oxide (˙NO), carbonate radical-anion (CO 3 ˙- ), peroxynitrite (ONOO - ), and superoxide (O 2 ˙- ) were increased in oral cancer cells following manoalide treatments in terms of fluorescence staining and flow cytometry. Cellular reactive species (˙OH, NO 2 · , ˙NO, CO 3 ˙- , and ONOO - ) as well as cellular and mitochondrial reactive species (O 2 ˙- ) were induced in oral cancer cells following manoalide treatment for 6 h. NAC, MitoTEMPO, and ZVAD inhibit manoalide-induced apoptosis in terms of annexin V and pancaspase activity assays. Moreover, NAC inhibits mitochondrial reactive species and MitoTEMPO inhibits cellular reactive species, suggesting that cellular and mitochondrial reactive species can crosstalk to regulate each other. ZVAD shows suppressing effects on the generation of both cellular and mitochondrial reactive species. In conclusion, manoalide induces reciprocally activation between cellular and mitochondrial reactive species and apoptosis in oral cancer cells., Competing Interests: The authors declare no conflict of interest., (Copyright © 2021 Hui-Ru Wang et al.)

Withaferin A (WFA), a Withania somnifera -derived triterpenoid, is an anticancer natural product. The anticancer effect of nonionizing radiation such as ultraviolet-C (UVC) as well as the combined treatment of UVC and WFA is rarely investigated. Low dose UVC and/or WFA treatments (12 J/m 2 and/or 1 μM) were chosen to evaluate antioral cancer cell line effects by examining cytotoxicity, cell cycle disruption, apoptosis induction, and DNA damage. For two cancer cell lines (Ca9-22 and HSC-3), single treatment (UVC or WFA) showed about 80% viability, while a combined treatment of UVC/WFA showed about 40% viability. In contrast, there was noncytotoxicity to normal oral cell lines (HGF-1). Compared to single treatment and control, low dose UVC/WFA shows high inductions of apoptosis in terms of flow cytometric detections for subG1, annexin V, pancaspase changes as well as Western blotting for detecting cleaved poly (ADP-ribose) polymerase (c-PARP) and caspase 3 (c-Cas 3) and luciferase assay for detecting Cas 3/7 activity. Low dose UVC/WFA also showed high inductions of oxidative stress and DNA damage in terms of flow cytometric detections of reactive oxygen species (ROS), mitochondrial superoxide (MitoSOX) generation, and membrane potential (MitoMP) destruction, γH2AX and 8-oxo-2'deoxyguanosine (8-oxodG) types of DNA damages. For comparison, low dose UVC/WFA show rare inductions of annexin V, Cas 3/7 activity, ROS, MitoSOX, and MitoMP changes to normal oral HGF-1 cells. Therefore, low dose UVC/WFA provides a novel selectively killing mechanism to oral cancer cells, suggesting that WFA is a UVC sensitizer to inhibit the proliferation of oral cancer cells.

The antibiotic antimycin A (AMA) is commonly used as an inhibitor for the electron transport chain but its application in anticancer studies is rare. Recently, the repurposing use of AMA in antiproliferation of several cancer cell types has been reported. However, it is rarely investigated in oral cancer cells. The purpose of this study is to investigate the selective antiproliferation ability of AMA treatment on oral cancer cells. Cell viability, flow cytometry, and western blotting were applied to explore its possible anticancer mechanism in terms of both concentration- and exposure time-effects. AMA shows the higher antiproliferation to two oral cancer CAL 27 and Ca9-22 cell lines than normal oral HGF-1 cell lines. Moreover, AMA induces the production of higher reactive oxygen species (ROS) levels and pan-caspase activation in oral cancer CAL 27 and Ca9-22 cells than in normal oral HGF-1 cells, providing the possible mechanism for its selective antiproliferation effect of AMA. In addition to ROS, AMA induces mitochondrial superoxide (MitoSOX) generation and depletes mitochondrial membrane potential (MitoMP). This further supports the AMA-induced oxidative stress changes in oral cancer CAL 27 and Ca9-22 cells. AMA also shows high expressions of annexin V in CAL 27 and Ca9-22 cells and cleaved forms of poly (ADP-ribose) polymerase (PARP), caspase 9, and caspase 3 in CAL 27 cells, supporting the apoptosis-inducing ability of AMA. Furthermore, AMA induces DNA damage (γH2AX and 8-oxo-2'-deoxyguanosine [8-oxodG]) in CAL 27 and Ca9-22 cells. Notably, the AMA-induced selective antiproliferation, oxidative stress, and DNA damage were partly prevented from N-acetylcysteine (NAC) pretreatments. Taken together, AMA selectively kills oral cancer cells in an oxidative stress-dependent mechanism involving apoptosis and DNA damage., (© 2020 Wiley Periodicals LLC.)

Some withanolides, particularly the family of steroidal lactones, show anticancer effects, but this is rarely reported for withanolide C (WHC)-especially anti-breast cancer effects. The subject of this study is to evaluate the ability of WHC to regulate the proliferation of breast cancer cells, using both time and concentration in treatment with WHC. In terms of ATP depletion, WHC induced more antiproliferation to three breast cancer cell lines, SKBR3, MCF7, and MDA-MB-231, than to normal breast M10 cell lines. SKBR3 and MCF7 cells showing higher sensitivity to WHC were used to explore the antiproliferation mechanism. Flow cytometric apoptosis analyses showed that subG1 phase and annexin V population were increased in breast cancer cells after WHC treatment. Western blotting showed that cleaved forms of the apoptotic proteins poly (ADP-ribose) polymerase (c-PARP) and cleaved caspase 3 (c-Cas 3) were increased in breast cancer cells. Flow cytometric oxidative stress analyses showed that WHC triggered reactive oxygen species (ROS) and mitochondrial superoxide (MitoSOX) production as well as glutathione depletion. In contrast, normal breast M10 cells showed lower levels of ROS and annexin V expression than breast cancer cells. Flow cytometric DNA damage analyses showed that WHC triggered γH2AX and 8-oxo-2'-deoxyguanosine (8-oxodG) expression in breast cancer cells. Moreover, N -acetylcysteine (NAC) pretreatment reverted oxidative stress-mediated ATP depletion, apoptosis, and DNA damage. Therefore, WHC kills breast cancer cells depending on oxidative stress-associated mechanisms.

Ultraviolet-C (UVC) irradiation provides an alternative radiotherapy to X-ray. UVC sensitizer from natural products may improve radiotherapy at low cytotoxic side effects. The aim of this study is to assess the regulation for oral cancer cell proliferation by a combined treatment of UVC and our previously reported anti-oral cancer natural product (ethyl acetate extract of Nepenthes adrianii × clipeata ; EANA). The detailed possible UVC sensitizing mechanisms of EANA such as effects on cell proliferation, cell cycle, apoptosis, and DNA damage are investigated individually and in combination using 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTS) assay, flow cytometry, and western blotting at low dose conditions. In a 24 h MTS assay, the low dose EANA (5 μg/mL) and low dose UVC (12 J/m 2 ) individually show 80% and combinedly 57% cell proliferation in oral cancer Ca9-22 cells; but no cytotoxicity to normal oral HGF-1 cells. Mechanistically, low dose EANA and low dose UVC individually induce apoptosis (subG1 accumulation, pancaspase activation, and caspases 3, 8, 9), oxidative stress (reactive oxygen species, mitochondrial superoxide, and mitochondrial membrane potential depletion), and DNA damage (γH2AX and 8-hydroxy-2'-deoxyguanosine). Moreover, the combined treatment (UVC/EANA) synergistically induces these changes. Combined low dose treatment-induced antiproliferation, apoptosis, oxidative stress, and DNA damage were suppressed by the ROS scavenger N -acetylcysteine. In conclusion, UVC/EANA shows synergistic antiproliferation, oxidative stress, apoptosis, and DNA damage to oral cancer cells in an oxidative stress-dependent manner. With the selective killing properties of low dose EANA and low dose UVC, EANA provides a novel UVC sensitizing agent to improve the anti-oral cancer therapy.

The sensitizing effect of chromone-derived compounds on UVC-induced proliferation inhibition has not been comprehensively investigated so far. The subject of this study was to examine the proliferation change of oral cancer cells while using the combined treatment of UVC (254 nm) with our previously developed sulfonyl chromen-4-ones (CHW09), namely UVC/CHW09. Cell viability, apoptosis, oxidative stress, and DNA damage for the individual and combined treatments for UVC and/or CHW09 were examined in oral cancer Ca9-22 cells. In 24 h MTS assay, UVC (30 J/m 2 ; UVC30), or CHW09 (25 and 50 µg/mL; namely, CHW09-25 and CHW09-50) show 54%, 59%, and 45% viability. The combined treatment (UVC30/CHW09-25 and UVC30/CHW09-50) show lower cell viability (45% and 35%). Mechanistically, UVC/CHW09 induced higher apoptosis than individual treatments and untreated control, which were supported by the evidence of flow cytometry for subG1, annexin V/7-aminoactinomycin D, pancaspase and caspases 3/7 activity, and western blotting for cleaved poly(ADP-ribose) polymerase. Moreover, this cleaved PARP expression was downregulated by pancaspase inhibitor Z-VAD-FMK. UVC/CHW09 showed higher oxidative stress than individual treatments and untreated control in terms of flow cytometry for reactive oxygen species, mitochondrial membrane potential, and mitochondrial mass. Furthermore, UVC/CHW09 showed higher DNA damage than individual treatments and untreated control in terms of flow cytometry for H2A histone family member X and 8-oxo-2'-deoxyguanosine. In conclusion, combined treatment UVC/CHW09 suppresses proliferation, and promotes apoptosis, oxidative stress, and DNA damage against oral cancer cells, providing a novel application of sulfonyl chromen-4-ones in order to sensitize UVC induced proliferation inhibition for oral cancer therapy.

Some lichens provide the resources of common traditional medicines and show anticancer effects. However, the anticancer effect of Usnproliea barbata ( U. barbata ) is rarely investigated, especially for oral cancer cells. The aim of this study was to investigate the cell killing function of methanol extracts of U. barbata (MEUB) against oral cancer cells. MEUB shows preferential killing against a number of oral cancer cell lines (Ca9-22, OECM-1, CAL 27, HSC3, and SCC9) but rarely affects normal oral cell lines (HGF-1). Ca9-22 and OECM-1 cells display the highest sensitivity to MEUB and were chosen for concentration effect and time course experiments to address its cytotoxic mechanisms. MEUB induces apoptosis of oral cancer cells in terms of the findings from flow cytometric assays and Western blotting, such as subG1 accumulation, annexin V detection, and pancaspase activation as well as poly (ADP-ribose) polymerase (PARP) cleavage. MEUB induces oxidative stress and DNA damage of oral cancer cells following flow cytometric assays, such as reactive oxygen species (ROS)/mitochondrial superoxide (MitoSOX) production, mitochondrial membrane potential (MMP) depletion as well as overexpression of γH2AX and 8-oxo-2'deoxyguanosine (8-oxodG). All MEUB-induced changes in oral cancer cells were triggered by oxidative stress which was validated by pretreatment with antioxidant N -acetylcysteine (NAC). In conclusion, MEUB causes preferential killing of oral cancer cells and is associated with oxidative stress, apoptosis, and DNA damage.

Objectives: In current guidelines, early tongue cancer status post partial glossectomy without adverse risk features do not require adjuvant treatment. However, many of these patients developed recurrence with neck metastases soon. The objectives of this study were to investigate the potential risk factors in early tongue cancer that prophylactic management of neck may be considered., Materials and Methods: From January 2010 to September 2015, this retrospective study enrolled 102 patients with T1-2N0 primary oral tongue squamous cell carcinoma according to AJCC 8th edition Cancer Staging System. All patients underwent partial glossectomy with or without selective neck dissection, and did not receive any adjuvant treatment. Patients with any adverse risk features were excluded. We have studied the 4-year cancer-specific survival and neck recurrence rate, and analyzed the relevance between pathologic tumor classification, tumor depth, tumor histologic grade, and measured surgical margin of primary tumor., Results: The median follow up duration was 47 months (range 6-93 months) with the median recurrence interval was 13 months. Histologic grade ≥2 of primary tumor was significantly associated with increased risk of neck recurrence and disease-specific mortality in both univariate and multivariate analysis., Conclusion: Histologic grade ≥2 was an adverse prognostic factor of neck recurrence and was significantly associated with poor cancer-specific survival in T1-2N0 early oral tongue cancer patients. Therefore, prophylactic neck dissection or prophylactic adjuvant radiation therapy to neck may be considered in T1-2N0 early oral tongue cancer with histologic grade ≥2 of primary tumor., Competing Interests: Declaration of Competing Interest The author declare that there is no conflict of interest., (Copyright © 2020 Elsevier Ltd. All rights reserved.)