Combination Of Autophagy Inhibitors Research Articles

Active fraction of Polyrhachis vicina (Roger) alleviated cerebral ischemia/reperfusion injury by targeting SIRT3-mediated mitophagy and angiogenesis

BackgroundDamaged mitophagy and impaired angiogenesis involve in the pathogenic development of ischemic stroke. Active fraction of Polyrhachis vicina (Roger) (AFPR) showed great potential on neurological disease with it's remarkable anti-inflammatory and anti-oxidative effects. PurposeThis study designed to clarify the correlation between Pink1/Parkin-mediated mitophagy and angiogenesis after stroke, and to elucidate the role of SIRT3 in regulating mitophagy and angiogenesis, and to address the mechanism of AFPR on promoting mitophagy and angiogenesis in microvessels endothelium of ischemic brain. Study designA cerebral ischemia/reperfusion (CIR) rat model was developed by middle cerebral artery occlusion procedure. bEnd.3 cells were exposed to oxygen-glucose deprivation/reoxygenation (OGD/R) to mimic CIR process. Neurological function, mitophagy and angiogenesis related indicators were measured. SIRT3 siRNA and 3-MA were used to verify the interaction between SIRT3-mediated mitophagy and angiogenesis. MethodsCIR rats were orally treated with AFPR (8 and 4 g raw drug /kg) and Nimodipine (10.8 mg/kg) for 12 days to mimic the recovery phase post-stroke. The neurological function assessment, TTC staining, HE staining, TUNEL staining and Nissl staining were performed to assess neuroprotective effects of AFPR against CIR. Then CD31-labeled microvessel density in brain was visualized and quantified by immunofluorescence staining. Mitochondrial ultrastructure was assessed by transmission electron microscope scanning. Expressions of relative proteins,e.g. SIRT3, Pink1, Parkin, LC3-II, p62, VEGFA, involving in mitophagy and angiogenesis, were detected by Western blotting analysis. In vitro, bEnd.3 cells were cultured with AFPR or in combination of autophagy inhibitor 3-MA during the reoxygenation. Then cell viability, and LDH releasing were measured. Angiogenic indicators,such as migration and tube formation activity, VEGFA level were determined. To assess effects of AFPR on mitophagy, mitophagy-related proteins were detected, as well as the autophagosome engulfment and lysosome degradation of mitochondria. To address the role of SIRT3, deacetylation activity of SIRT3 was validated by detecting acetylated FOXO3A level with co-immunoprecipitation (Co-IP) assay. Pre-treatment of siRNA or combination use of 3-MA were used to verify the detailed mechanism. ResultsAFPR remarkably reduced neurological scores and infarct size, alleviated neuron apoptosis in cortex, and increased Nissl density in hippocampus of CIR rats. In addition, AFPR significantly promoted angiogenesis by increasing microvessels density and VEGFA expressions, increased SIRT3 expression, and activated Pink1/Parkin mediated mitophagy. In bEnd.3 cells, the combination use of 3-MA and AFPR further demonstrated that AFPR might promote angiogenesis after OGD/R injury through activating Pink1/Parkin mediated mitophagy. Co-IP assay suggested AFPR reduced acetylated FOXO3A level. This might be correlated with an elevation of SIRT3 expression and it's deacetylation activity. SIRT3 siRNA pretreatment significantly abolished the activation of mitophagy through Pink1/Parkin axis, eventually inhibited angiogenesis. ConclusionAFPR promoted angiogenesis through activating mitophagy after cerebral ischemia reperfusion, which might partially involved in the amelioration of SIRT3-mediated regulation on Pink1/Parkin axis. Our study will shed new light on the role of SIRT3 in ischemic brain, especially in regulating mitophagy and angiogenesis after stroke.

Temozolomide and chloroquine co-loaded mesoporous silica nanoparticles are effective against glioma

The past decades have witnessed great progress in nanoparticle‐based cancer‐targeting drug delivery systems, but their therapeutic potentials is yet to be fully exploited. In this research, temozolomide (TMZ) and chloroquine (CQ) were loaded into the mesoporous silica nanoparticles (MSNs), the surface was coated with polydopamine (PDA), and the complex was coupled with arginine-glycine-aspartic (RGD) to successfully prepare TMZ/CQ@MSN-RGD. RGD-MSNs accumulated more in the cell and tumor models than in unmodified MSNs in the in vitro and in vivo experiments and can directly induce apoptosis and autophagy in tumor cells. In addition, TMZ/CQ@MSN-RGD therapy enhanced the apoptosis effect of the RGD-MSNs in glioma. Therefore, the combination of autophagy inhibitor with chemotherapy drugs in nanocarriers may promote therapeutic efficacy in treating glioma.

Effect of Autophagy Inhibitors on Radiosensitivity in DNA Repair-Proficient and -Deficient Glioma Cells.

Background and Objectives: The development of radioresistance is a fundamental barrier to successful glioblastoma therapy. Autophagy is thought to play a role in facilitating the DNA repair of DNA damage foci in radiation-exposed tumor cells, thus, potentially contributing to their restoration of proliferative capacity and development of resistance in vitro. However, the effect of autophagy inhibitors on DNA damage repair is not fully clear and requires further investigation. Materials and Methods: In this work, we utilized M059K (DNA-PKcs proficient) and M059J (DNA-PKcs deficient) glioma cell lines to investigate the role of autophagy inhibitors in the DNA repair of radiation-induced DNA damage. Cell viability following radiation was determined by trypan blue exclusion in both cell lines. Cell death and autophagy assays were performed to evaluate radiation-induced cell stress responses. DNA damage was measured as based on the intensity of phosphorylated γ-H2AX, a DNA double-stranded breaks (DSBs) marker, in the presence or absence of autophagy inhibitors. Results: The cell viability assay showed that M059J cells were more sensitive to the same dose of radiation (4 Gy) than M059K cells. This observation was accompanied by an elevation in γ-H2AX formation in M059J but not in M059K cells. In addition, the DAPI/TUNEL and Senescence-associated β-galactosidase (SA-β-gal) staining assays did not reveal significant differences in apoptosis and/or senescence induction in response to radiation, respectively, in either cell line. However, acridine orange staining demonstrated clear promotion of acidic vesicular organelles (AVOs) in both cell lines in response to 4 Gy radiation. Moreover, DNA damage marker levels were found to be elevated 72 h post-radiation when autophagy was inhibited by the lysosomotropic agent bafilomycin A1 (BafA1) or the PI3K inhibitor 3-methyl adenine (3-MA) in M059K cells. Conclusions: The extent of the DNA damage response remained high in the DNA-PKcs deficient cells following exposure to radiation, indicating their inability to repair the newly formed DNA-DSBs. On the other hand, radioresistant M059K cells showed more DNA damage response only when autophagy inhibitors were used with radiation, suggesting that the combination of autophagy inhibitors with radiation may interfere with DNA repair efficiency.

Synergistic effects of autophagy inhibitors combined with cisplatin against cisplatin-resistant nasopharyngeal cancer cells.

This study explored the synergistic effects of autophagy inhibitors combined with cisplatin against cisplatin-resistant nasopharyngeal cancer cells by treating HNE-1 and cisplatin (diamminedichloroplatinum; DDP)-resistant HNE1/DDP nasopharyngeal cancer cell lines with DDP, autophagy inhibitors, or a combination of autophagy inhibitors and DDP. Cell viability was determined via MTT (colorimetric) and colony-forming assays, and the rate of apoptosis was determined using propidium iodide (PI) and annexin V double-staining. The expressions of proteins were determined by Western blotting. For our in-vivo studies, a murine xenograft model was established to evaluate the anti-tumor effects of the combination of autophagy inhibitor and DDP. The results showed that treatment with DDP increased the expressions of ATP-binding cassette sub-family B member 1 (ABCB1), ATP Binding Cassette Subfamily C Member 1 (ABCC1), and P-glycoprotein 1 (P-gp) in the HNE1/DDP cell lines. Treatment with chloroquine decreased the expression levels of ABCB1, ABCC1, and P-gp, and increased the formation of LC3-II and the expression levels of p62 in the HNE1/DDP cells. Additionally, the combination of autophagy inhibitors and DDP produced a synergistic effect on DDP-induced cell death and apoptosis. Furthermore, the combination of the autophagy inhibitor and DDP showed significant anti-tumor effects in the xenograft mouse model. In summary, autophagy inhibitors show synergistic anti-tumor effects with DDP in vitro against DDP-resistant nasopharyngeal cancer cells and in vivo in our xenograft murine model.

A promising therapeutic combination for metastatic prostate cancer: Chloroquine as autophagy inhibitor and palladium(II) barbiturate complex

Autophagy is a catabolic process for cells that can provide energy sources and allows cancer cells to evade cell death. Therefore, studies on the combination of autophagy inhibitors with drugs are increasing as a new treatment modality in cancer. Previously, we reported the anti-tumor activity of a Palladium (Pd)(II) complex against different types of cancer in vitro and in vivo. Chloroquine (CQ), the worldwide used anti-malarial drug, has recently been focused as a chemosensitizer in cancer treatment. The aim of this study was to investigate the efficacy of a combined treatment of these agents that work through different mechanisms to provide an effective treatment modality for metastatic prostate cancer that is certainly fatal. Metastatic prostate cancer cell lines (PC-3 and LNCaP) were treated with Pd (II) complex, CQ, and their combination. The combination enhanced apoptosis by increasing phosphatidylserine translocation and pro-apoptotic proteins. Apoptosis was confirmed by the use of apoptosis inhibitor. The formation of acidic vesicular organelles (AVOs) was observed by acridine orange staining in fluorescence microscopy. The Pd (II) complex increased AVOs formation in prostate cancer cells and CQ-pretreatment has potentiated this effect. Importantly, treatment with CQ suppressed the pro-survival function of autophagy, which might have contributed to enhanced cytotoxicity. In addition, PI3K/AKT/mTOR-related protein expressions were altered after the combination of treatments. Our results suggest that combination treatment enhances apoptotic cell death possibly via the inhibition of autophagy, and may therefore be regarded as a novel and better approach for the treatment of metastatic prostate cancer.

Autophagy and Tumour Stem Cells.

Autophagy is critical for the survival and stemness maintenance of cancer stem cells (CSCs) and is an enhancer of CSC tumorigenesis. At the same time, autophagy contributes to conditions optimal for facilitating the invasion and metastasis of CSCs. Moreover, autophagy induces the dormant state of CSCs to help them resist the cytotoxic effects of chemotherapy andradiotherapy, thereby improving the likelihood of their survival. The combination of autophagy inhibitors with specific drugs targeting specific CSC subpopulations is expected to act specifically on CSCs and produce fewer toxic side effects on normal tissues. This in-depth study is very timely and important for further identifying the potential role of autophagy in different states of CSCs and places a particular emphasis on exploring molecular mechanisms in the regulation of autophagy via advanced techniques based on molecular biology.

Abstract 4273: Inhibition of autophagy increases HNSCC sensitivity to cancer therapies

Abstract Background: Radiation and EGFR-targeted therapies are commonly used in the treatment of head and neck squamous cell carcinoma (HNSCC). These treatments fail to control a significant number of cancers resulting in a 5-year survival rate that remains around 40-50%. We have shown that both cetuximab and radiation induce autophagy, a pro-survival cellular stress response, in head and neck cancer. In this study, we examine the consequence of autophagy inhibition and investigate the molecular mechanism underlying therapy-induced autophagy. Methods: Autophagy was assessed using a nano-Luc LC3 reporter (Promega), immunofluorescence for LC3, p62, and acridine orange in HNSCC cell lines. RNAi knockdown of EGFR and LAPTM4B were used to test the involved signaling molecules. Radiation was delivered using a RS225 cabinet irradiator at a dose rate of approximately 3 Gy/min with dose validation by TLD using custom phantoms. Cetuximab was delivered via intraperitoneal injection. Vps34 inhibitor SAR405 and ULK1 inhibitor SBI-0206965 were used to determine whether inhibition of autophagy reduces cell survival or represses cancer cell growth in the clonogenic assay. A flank xenograft model using A253 cells was used to test the combination of autophagy inhibitors and current therapies in vivo. Results: As previously shown, both cetuximab and radiation induced autophagy by two times. Knockdown of EGFR and LAPTM4B decreased autophagy (62.5% and 65%, respectively) when assessed using the nano-Luc reporter assay. Similar results were seen using IF. Using a clonogenic survival assay, the combination of SAR405 and radiation resulted in complete loss of cell survival suggesting a radiosensitizing effect. In vivo, SAR405 treatment improved tumor control when combined with radiation or cetuximab when compared to either treatment alone. Conclusions: Therapy induced autophagy is dependent upon expression of both EGFR and LAPTM4B. Inhibition of autophagy resulted decreased cell survival in vitro and resulted in decreased in vivo tumor growth. These results suggest that inhibition of autophagy may be a viable approach to sensitize HNSCC to anti-cancer treatments. Citation Format: Yong-Syu Lee, Justin Skiba, Jaimee Kubatzke, Kwangok P. Nickel, Randall Kimple. Inhibition of autophagy increases HNSCC sensitivity to cancer therapies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4273.

Sodium butyrate induces autophagic apoptosis of nasopharyngeal carcinoma cells by inhibiting AKT/mTOR signaling

Previously, we confirmed the anti-tumor effects of sodium butyrate (NaBu) in nasopharyngeal carcinoma (NPC). However, its molecular mechanisms have not be fully elucidated. In this study, we studied the effects of NaBu on autophagy and explored the relation between NaBu associated autophagy and apoptosis in NPC cells. EGFP-LC3 plasmids were introduced into NPC cells to observed the effects of NaBu on autophagy flux with or without chloroquine (CQ) addition. Autophagy markers were also detected by Western blot. Under NaBu treatment, autophagy and apoptosis markers were detected simultaneously at different time. Then, to explore the roles of autophagy in NaBu induced apoptosis, the effects of autophagy inhibition, via specific inhibitor treatment or key gene knockdown, were analyzed. At last, the upstream signaling and its roles in NaBu induced autophagy and apoptosis were also analyzed. Increased LC3 dots and LC3-II accumulation indicated that NaBu can promote autophagy flux in NPC cells. LC3-II accumulation was earlier than cleaved PARP increment suggesting autophagy activation is prior to apoptosis activation, which was validated by flow cytometry mediated apoptosis analysis. Moreover, autophagy inhibition, achieved by 3-MA treatment or BECN1 knockdown, can antagonize NaBu induced apoptosis reflecting by re-deregulated cPARP and apoptotic rates. Furthermore, NaBu treatment inhibited the AKT/mTOR axis indicated by deregulated p-AKT(S473) and p-mTOR(S2448) and ectopic AKT expression both suppressed NaBu induced autophagy and apoptosis. At last, Western blot showed that HDAC6 dependent EGFR deregulation may account for the NaBu associated AKT/mTOR inhibition. NaBu can induce autophagic apoptosis via suppressing AKT/mTOR axis in NPC cells. Our results suggest that combination of autophagy inhibitors and deacetylase inhibitors may not be recommended in NPC clinical treatment.

Novel Ridaifen-B Structure Analog Induces Apoptosis and Autophagy Depending on Pyrrolidine Side Chain.

Ridaifen (RID)-B is an analog derived from tamoxifen (TAM). TAM has an antitumor effect by acting as an antagonist to estrogen receptor (ER). However, TAM is known to also induces apoptosis in cancer cells that do not have ER. We clarified that RID-B induces cell death at a lower concentration than TAM, and causes ER-independent apoptosis and autophagy. Based on the results of previous studies, we assumed that RID-B had a unique target different from ER and examined structural activity correlation to determine what kinds of structural features are related to RID-B activity. As a result, we found there was activity even without one of phenyl groups (Ar3) in RID-B and revealed that two pyrrolidine side chains peculiar to RID-B are related to the action. Furthermore, analogs with shorter alkyl side chains induced autophagy, but analogs with certain length of alkyl side chains induced apoptosis. Also, although there is no doubt that RID-B induces apoptosis by causing mitochondrial injury, our results suggested that such injury induced mitochondria-selective autophagy. We revealed that RID-B induce mitophagy and that this mitophagy is a defense mechanism against RID-B. Our results suggest that autophagy was induced against apoptosis caused by mitochondrial dysfunction in RID-B, so the combination of autophagy inhibitor and anticancer-drug can be effective for cancer treatment.

Divaricoside Exerts Antitumor Effects, in Part, by Modulating Mcl-1 in Human Oral Squamous Cell Carcinoma Cells

Cardiac glycosides (CGs), prescribed to treat congestive heart failure and arrhythmias, exert potent antitumor activity. In this study, divaricoside (DIV), a CG isolated from Strophanthus divaricatus was examined for its antitumor potency in oral squamous cell carcinoma (OSCC) cells. Cell growth was inhibited by DIV in a dose- and time-dependent manner in SCC2095 and OECM-1 OSCC cells using MTT assays. DIV induced S and G2/M phase arrest accompanied by downregulation of phosphorylated CDC25C, CDC25C, and CDC2 in SCC2095 cells. In addition, DIV induced apoptosis by activating caspase-3 and downregulating the expression of Mcl-1. Furthermore, overexpression of Mcl-1 partially reversed DIV-induced death in SCC2095 cells. Additionally, western blot and transmission electron microscopy analyses also indicated that DIV induced autophagy in SCC2095 cells. However, the combination of autophagy inhibitor did not affect DIV-mediated apoptosis in SCC2095 cells. Together, these findings suggest that translational potential of DIV to be developed as a therapeutic agent for OSCC treatment.

Autophagy blockade sensitizes human head and neck squamous cell carcinoma towards CYT997 through enhancing excessively high reactive oxygen species-induced apoptosis.

• CYT997 exhibits anticancer activity by induction of ROS-associated apoptosis. • mTOR-dependent cytoprotective autophagy prevents CYT997-induced apoptosis. • Blockade of autophagy augments CYT997 efficacy by enhanced ROS-associated apoptosis. • Combination of autophagy inhibitors with CYT997 is more effective against HNSCC.

Effect and mechanism of Stellettin B induced human glioblastoma SF295 cells autophagy

Objective To study the autophagy of human glioblastoma SF295 cells induced by sponge-derived triterpenoid Stellittin B (Stel B) and to discuss the related mechanism. Methods The proliferation inhibitory activity of Stel B on SF295 cells was studied by WST-8 assay. The autophagy effect induced by Stel B on SF295 cells was evaluated bya variety of experimental techniques, including MDC staining, Western Blot assay, and mRFP-GFP-LC3 plasmid transfection method.The activity of Stel B on the representative signal proteins in PI3K/Akt/mTOR pathway in SF295 cells was examined by Western Blot. The anti-tumor activity of Stel B was detected by WST-8 assay after blocking autophagy by autophagy inhibitor chloroquine. Results Stel B could significantly inhibit the proliferation of SF295 cells with IC50 value as 0.026 μmol/L. Plenty of autophagosome was found in Stel B treated A549 cells by MDC staining. The expression of autophagy marker proteins including LC3B-II increased. The confocal microscopy results showed that Stel B promoted autophagosome forming and inhibitedthe fusion of autophagosome with lysosome. The Western Blot results showed that Stel B inhibited the expression of PI3K-p110 protein in SF295 cells and decreased the phosphorylation level of Akt and mTOR. The inhibitory effects of Stel B with different concentrations on SF295 cells were all enhanced by the inhibition of autophagy with chloroquine, and the differences were statistically significant(all P<0.05). Conclusion Stel B can induce SF295 cells autophagy via blocking PI3K/Akt/mTOR pathway. This autophagy effect is beneficial to the survival of SF295 cells. The antitumor activity of Stel B can be enhanced by a combination of autophagy inhibitors. Key words: Stellettin B; Human glioblastoma; Autophagy; SF295 cell; PI3K/Akt/mTOR pathway

Abstract 449: Reduced argininosuccinate synthetase expression in refractory sarcomas: impacts on therapeutic potential and drug resistance

Abstract Introduction: Treating drug-resistant sarcomas remain a major challenge. The present study aimed to identify a novel therapy for drug-resistant sarcomas based on a metabolic errors involving argininosuccinate synthetase1 (ASS1). Methods: We assessed the expression of ASS1 and P-glycoprotein (P-gp) in osteosarcoma (KHOS), doxorubicin (Dox)-resistant osteosarcoma (KHOSR2), epithelioid sarcomas (ES-X and VAESBJ), alveolar soft part sarcoma (ASPS-KY), and each clinical specimen. Each cell was cultured in arginine-containing and arginine-free media. Cell growth was assessed using an XTT assay and flow cytometry. We analyzed the induction of autophagy in arginine-free medium. Moreover, we assessed the expression of P-gp in after suppressing ASS1 in Dox-sensitive cells (MCF-7, KHOS) and in after transfecting the ASS1 into Dox-resistant cells (ES-X, VAESBJ, ASPS-KY and KHOSR2). Results: The expression of ASS1 was reduced in Dox-resistant sarcoma cells. Immunohistochemistry (IHC) and real-time PCR showed that there was interestingly an inverse relationship between the expression of ASS1 and the expression of P-gp. The inhibition of cellular proliferation with G1-arrest was shown to lead to autophagy with arginine deprivation. In addition, the combination of autophagy inhibitor plus arginine deprivation was more effective than arginine deprivation alone. In cells in which the expression of ASS1 was suppressed, the expression of P-gp was upregulated in comparison to negative controls. Discussion: These results indicate that the reduced expression of ASS1 expression in Dox-resistant sarcomas may contribute to drug resistance. ASS1 deficiency is a potential target for novel drug therapies. The combination of arginine deprivation therapy and an autophagy inhibitor may have anti-tumor effects in refractory sarcomas. Significance: As the induction of autophagy by the deprivation arginine may have a pro-survival role in patients with ASS1-deficient sarcomas, the combination of arginine deprivation therapy with autophagy modulators might potentiate anti-tumor effects in patients with drug-resistant sarcomas. Citation Format: Eisuke Kobayashi, Youngji Kim, Daisuke Kubota, Yoshiyuki Suehara, Akira Kawai, Shigehisa Kitano. Reduced argininosuccinate synthetase expression in refractory sarcomas: impacts on therapeutic potential and drug resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 449. doi:10.1158/1538-7445.AM2017-449

Knockdown of retinoblastoma protein may sensitize glioma cells to cisplatin through inhibition of autophagy

Glioblastoma multiforme (GBM) is one of the deadliest forms of cancer due to its limited sensitivity to chemotherapy and radiotherapy. Cisplatin (CCDP) is a widely used chemotherapeutic agent for tumors, but the agent often results in the development of chemo-resistance. In several cancers, cisplatin resistance is associated with autophagy induction. Here, we found that in glioma cells cisplatin treatment induced autophagy. Our data indicates that the autophagy induction plays a critical role in cisplatin resistance of glioma cells, knockdown of RB inhibited autophagy induced by cisplatin, and inhibition of autophagy improved cisplatin-induced apoptosis. It suggests that a combination of autophagy inhibitors with cisplatin may improve the therapeutic efficiency of cisplatin towards GBM with acquired resistance.

Inhibition of autophagy by 3-MA enhances IL-24-induced apoptosis in human oral squamous cell carcinoma cells.

BackgroundInterleukin-24(IL-24), also referred to as melanoma differentiation-associated gene-7(mda-7), is a unique member of the IL-10 gene family, which displays nearly ubiquitous cancer-specific toxicity. The most notable feature of IL-24 is selectively induced growth suppression and apoptosis in various cancer cells, with no harmful effects toward normal cells. Autophagy is a self-protective mechanism in many kinds of tumor cells that respond to anticancer treatment. It is reported that autophagy inhibition could enhance the effects of many kinds of anticancer treatments, including gene therapy. However, whether IL-24 is effective to treat oral squamous cell carcinomas (OSCC) and if autophagy inhibition could improve the anticancer effect of IL-24 towards OSCC is has not been detected.MethodsMTT assays were carried out to determine the cell proliferation; Transfection was used to gene transfer; Western Blot was performed to detect the protein level of LC3II, P62, Beclin 1, Cleaved caspase-3, β-Tubulin and β-actin; Apoptosis rates and cell cycle alteration were analyzed using flow cytometry; Autophagy induction was confirmed by MDC staining, GFP-LC3 staining and transmission electron microscopy. Amount of IL-24 in the culture medium was quantified by ELISA. Apoptosis in vivo was analyzed by TUNEL assay. HE staining was used to observe the morphology of the samples.ResultsIn the present study, we proved that IL-24 have a novel anticancer effect towards KB cells and that autophagy inhibition could improve the anticancer effect of IL-24. IL-24 treated cells showed autophagy characteristics and autophagy inhibition by 3-methyladenine (3-MA) significantly enhanced IL-24-induced apoptosis. Similar results were obtained in the KB cells xenograft tumor model.ConclusionsThese results suggest that the combination of autophagy inhibitors and IL-24 based on the AdLTR2EF1α-mediated gene transfer could be a promising way to cure OSCC.

Autophagy in Cancer Therapy: Progress and Issues

Autophagy is an evolutionarily conserved intracellular self-digestion process, which mediates homeostasis in response to various stresses via degradation of damaged organelles or unnecessary proteins. It has been demonstrated that autophagy involves in tumorigenesis and progression. Autophagy serves either as tumor suppressor or promotor in a context-dependent way. It has been revealed in multiple studies that autophagy plays a pro-survival role upon treatment of anticancer drugs. Thus, combination of autophagy inhibitors with anticancer drugs may provide a desirable strategy to improve therapeutic efficacy. In this review, we summarize recent progress in the process and regulation of autophagy with a highlight in advances in the role of autophagy in cancer treatment. We also summarize some recent clinical outcomes of combinatorial use of autophagy inhibitors and anticancer drugs, and introduce latest discovered selective autophagy inhibitors. Some issues which should be paid attention to during the research to improve the clinical outcomes are discussed.

Induction of autophagy by cisplatin‐resistant esophageal cancer cells promotes survival and recovery

Esophageal squamous cell carcinoma (ESCC) is a highly malignant disease with a poor prognosis. Cisplatin‐chemotherapy is widely used for the treatment of this malignancy. However, the clinical efficacy of this drug is limited due to intrinsic and acquired resistance to it. Autophagy is an evolutionarily conserved lysosomal degradation pathway. Induction of autophgay has emerged as a drug resistance mechanism that promotes cancer cell survival. In this study, we investigated cellular response after treatment with cisplatin in ESCC cells. Cisplatin‐resistanct cell lines exhibited reduced apoptosis and senescence compared with cisplatin‐sensitive counterparts. Moreover, cisplatin treatment induced autophagy in resistant cells but not in their sensitive counterparts. The induction of autophagy was evidenced by the formation of LC3+ autophagosomes and decreased p62 protein level and was accompanied by the suppression of mammalian target of rapamycin complex 1 activity. Inhibition of autophagy induction by knockdown of ATG5 or ATG7 significantly reduced the clonogenic survival of cisplatin‐treated resistant cells. Disruption of lysosomal activity with chloroquine improved chemotherapeutic effect of cisplatin in resistant cancer cells both in vitro and in vivo. In addition, cisplatin increased phosphorylation of ERK, p38 and JNK, but only inhibition of ERK activation significantly attenuated the recovery of cisplatin‐treated cells. Moreover, MEK inhibitor attenuated cisplatin‐induced cellular apoptosis and senescence. To conclude, this study demonstrates that an autophagic response to cisplatin is a survival mechanism that promotes chemo‐resistance and recovery. This finding suggests that the combination of autophagy inhibitors with cisplatin may have significant potential clinical application in the circumvention of cisplatin‐resistant ESCC.

Necroptosis modulated by autophagy is a predominant form of melanoma cell death induced by sanguilutine

We show that the plant quaternary benzo[c]phenanthridine alkaloid sanguilutine (SL) is a strong inducer of caspase-independent non-apoptotic death in human melanoma cells. Necrostatin-1, a specific inhibitor of necroptosis, completely reversed the cytotoxic effect of SL, suggesting that necroptosis was a predominant type of cell death induced by SL in these cells. In addition, we showed that SL can trigger an autophagic response, as confirmed by GFP-LC3 puncta formation and LC3-II accumulation. Interestingly, we observed a significant decrease in the viability of melanoma cells treated with combination of autophagy inhibitors (3-methyladenine, bafilomycin-A1 and LY294002) and SL. Our results further indicated that autophagy may serve as a pro-survival mechanism, delaying the induction of necroptosis in melanoma cells. The ability of SL to induce caspase-independent non-apoptotic cell death (necroptosis) suggests its possible therapeutic potential in the treatment of apoptosis-resistant melanoma tumours. Furthermore, SL might serve as a useful tool for studying the mechanisms of necroptosis and autophagy induction and the interplay between these two processes.