Berberine increases the killing effect of pirarubicin on HCC cells by inhibiting ATG4B-autophagy pathway.

Pirarubicin (THP) is a new generation of cell cycle non-specific anthracycline-based anticancer drug. In the clinic, THP and THP combination therapies have been shown to be effective in hepatocellular carcinoma (HCC) patients with transcatheter arterial chemoembolization (TACE) without serious side effects. However, drug resistance limits its therapeutic efficacy. Berberine (BBR), an isoquinoline alkaloid, has been shown to possess antitumour properties against various malignancies. However, the synergistic effect of BBR and THP in the treatment of HCC is unknown. In the present study, we demonstrated for the first time that BBR sensitized HCC cells to THP, including enhancing THP-induced growth inhibition and apoptosis of HCC cells. Moreover, we found that BBR sensitized THP by reducing the expression of autophagy-related 4B (ATG4B). Mechanistically, the inhibition of HIF1α-mediated ATG4B transcription by BBR ultimately led to attenuation of THP-induced cytoprotective autophagy, accompanied by enhanced growth inhibition and apoptosis in THP-treated HCC cells. Tumor-bearing experiments in nude mice showed that the combination treatment with BBR and THP significantly suppressed the growth of HCC xenografts. These results reveal that BBR is able to strengthen the killing effect of THP on HCC cells by repressing the ATG4B-autophagy pathway, which may provide novel insights into the improvement of chemotherapeutic efficacy of THP, and may be conducive to the further clinical application of THP in HCC treatment.

CircPHKB decreases the sensitivity of liver cancer cells to sorafenib via miR-1234-3p/CYP2W1 axis.

Sorafenib is currently the first-line treatment for patients with advanced liver cancer, but its therapeutic efficacy declines significantly after a few months of treatment. Therefore, it is of great importance to investigate the regulatory mechanisms of sorafenib sensitivity in liver cancer cells. In this study, we provided initial evidence demonstrating that circPHKB, a novel circRNA markedly overexpressed in sorafenib-treated liver cancer cells, attenuated the sensitivity of liver cancer cells to sorafenib. Mechanically, circPHKB sequestered miR-1234-3p, resulting in the up-regulation of cytochrome P450 family 2 subfamily W member 1 (CYP2W1), thereby reducing the killing effect of sorafenib on liver cancer cells. Moreover, knockdown of circPHKB sensitized liver cancer cells to sorafenib in vivo. The findings reveal a novel circPHKB/miR-1234-3p/CYP2W1 pathway that decreases the sensitivity of liver cancer cells to sorafenib, suggesting that circPHKB and the axis may serve as promising targets to improve the therapeutic efficacy of sorafenib against liver cancer.

Application of flipped classroom combined with virtual simulation platform in clinical biochemistry practical course.

BACKGROUND: The study explores an innovative teaching mode that integrates Icourse, DingTalk, and online experimental simulation platforms to provide online theoretical and experimental resources for clinical biochemistry practical courses. These platforms, combined with flipped classroom teaching, aim to increase student engagement and benefit in practical courses, ultimately improving the effectiveness of clinical biochemistry practical teaching. METHODS: In a prospective cohort study, we examined the impact of integrating the Icourse and DingTalk platforms to provide theoretical knowledge resources and clinical cases to 48 medical laboratory science students from the 2019 and 2020 grades. Students were assigned to the experimental group using an overall sampling method, and had access to relevant videos through Icourse before and during class. Using a flipped classroom approach, students actively participated in the design, analysis, and discussion of the experimental technique. For the experimental operation part, students participated in virtual simulation experiments and actual experiments. Overall, the study aimed to evaluate students' theoretical and operational performance after completing the practical course. To collect feedback, we distributed a questionnaire to students in the experimental group. For comparison, we included 42 students from the grades of 2017 and 2018 who received traditional instruction and were evaluated using standard textbooks as the control group. RESULTS: The experimental group scored significantly higher than the control group on both the theoretical and experimental operational tests (82.45 ± 3.76 vs. 76.36 ± 3.96, P = 0.0126; 92.03 ± 1.62 vs. 81.67 ± 4.19, P < 0.001). The survey revealed that the experimental group preferred the teaching mode that combined the flipped classroom with the virtual simulation platform. This mixed method effectively promoted understanding of basic knowledge (93.8%, 45/48), operative skills (89.6%, 43/48), learning interest (87.5%, 42/48), clinical thinking (85.4%, 41/48), self-learning ability (91.7%, 44/48), and overall satisfaction compared with traditional methods (P < 0.05). This study demonstrates that an innovative teaching approach significantly improves the quality of clinical biochemistry practical courses and promotes students' professional development and self-directed learning habits. CONCLUSION: Incorporating virtual simulation with flipped classrooms into clinical biochemistry practical teaching is an efficient and well-received alternative to traditional methods.

Dichloroacetate enhances the anti-tumor effect of sorafenib via modulating the ROS-JNK-Mcl-1 pathway in liver cancer cells.

Liver cancer is one of the most common and high recurrence malignancies. Besides radiotherapy and surgery, chemotherapy also plays an essential role in the treatment of liver cancer. Sorafenib and sorafenib-based combination therapies have been proven efficacy against tumors. However, previous clinical studies have indicated that some patients with liver cancer are resistant to sorafenib treatment and the existing strategies are not satisfactory in the clinic. Therefore, it is urgent to investigate strategies to improve the effectiveness of sorafenib for liver cancer and to explore effective drug combinations. In the present study, we found that dichloroacetate (DCA) could significantly enhance the anti-tumor effect of sorafenib on liver cancer cells, including reduced viability and dramatically promoted apoptosis in liver cancer cells. Moreover, compared to sorafenib alone, the combination of DCA and sorafenib markedly increased the degradation of anti-apoptotic protein Mcl-1 by enhancing its phosphorylation. Overexpression of Mcl-1 could significantly attenuate the synergetic effect of DCA and sorafenib on apoptosis induction in liver cancer cells. Furthermore, we found that the ROS-JNK pathway was obviously activated in the DCA combined sorafenib group. The levels of ROS and p-JNK were dramatically up-regulated in the two drug combination groups. Antioxidant NAC could alleviate the synergetic effects of DCA and sorafenib on ROS generation, JNK activation, Mcl-1 degradation, and cell apoptosis. Moreover, DCA and sorafenib's effects on Mcl-1 degradation and apoptosis could also be inhibited by JNK inhibitor 'SP'600125. Finally, the synergetic effects of DCA and sorafenib on tumor growth suppression, Mcl-1 degradation and induction of apoptosis were also validated in liver cancer xenograft in vivo. These findings indicate that DCA enhances the anti-tumor effect of sorafenib via the ROS-JNK-Mcl-1 pathway in liver cancer cells. This study may provide new insights to improve the chemotherapeutic effect of sorafenib, which may be beneficial for further clinical application of sorafenib in liver cancer treatment.

The PPARγ agonist rosiglitazone sensitizes the BH3 mimetic (-)-gossypol to induce apoptosis in cancer cells with high level of Bcl-2.

The BH3 mimetic (-)-gossypol (-)-G has shown promising efficacy to kill several kinds of cancer cells or potentiate current chemotherapeutics. But it induces limited apoptosis in cancer cells with high level of Bcl-2. The nuclear receptor PPARγ and its agonist rosiglitazone can suppress various malignancies. More importantly, rosiglitazone is able to enhance the anti-tumor effects of chemotherapy drugs such as carboplatin and tyrosine kinase inhibitors. In this study, we for the first time demonstrated that rosiglitazone could sensitize (-)-G to induce apoptosis in cancer cells with high level of Bcl-2. Furthermore, we found that (-)-G increased the mRNA level and protein stability of Mcl-1, which weakened the pro-apoptotic effect of (-)-G. Rosiglitazone attenuated the (-)-G-induced Mcl-1 stability through decreasing JNK phosphorylation. Additionally, rosiglitazone upregulated dual-specificity phosphatase 16 (DUSP16), leading to a reduction of (-)-G-triggered JNK phosphorylation. Animal experiments showed that rosiglitazone could sensitize (-)-G to repress the growth of cancer cells with high level of Bcl-2 in vivo. Taken together, our results suggest that the PPARγ agonists may enhance the therapeutic effect of BH3 mimetics in cancers with high level of Bcl-2 through regulating the DUSP16/JNK/Mcl-1 singling pathway. This study may provide novel insights into the cancer therapeutics based on the combination of PPARγ agonists and BH3 mimetics.

Hsp90 inhibitor 17-AAG sensitizes Bcl-2 inhibitor (-)-gossypol by suppressing ERK-mediated protective autophagy and Mcl-1 accumulation in hepatocellular carcinoma cells.

Natural BH3-memitic (-)-gossypol shows promising antitumor efficacy in several kinds of cancer. However, our previous studies have demonstrated that protective autophagy decreases the drug sensitivities of Bcl-2 inhibitors in hepatocellular carcinoma (HCC) cells. In the present study, we are the first to report that Hsp90 inhibitor 17-AAG enhanced (-)-gossypol-induced apoptosis via suppressing (-)-gossypol-triggered protective autophagy and Mcl-1 accumulation. The suppression effect of 17-AAG on autophagy was mediated by inhibiting ERK-mediated Bcl-2 phosphorylation while was not related to Beclin1 or LC3 protein instability. Meanwhile, 17-AAG downregulated (-)-gossypol-triggered Mcl-1 accumulation by suppressing Mcl-1(Thr163) phosphorylation and promoting protein degradation. Collectively, our study indicates that Hsp90 plays an important role in tumor maintenance and inhibition of Hsp90 may become a new strategy for sensitizing Bcl-2-targeted chemotherapies in HCC cells.

The Bcl-2/xL inhibitor ABT-263 increases the stability of Mcl-1 mRNA and protein in hepatocellular carcinoma cells.

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the major causes of mortality. ABT-263 is a newly synthesized, orally available Bcl-2/xL inhibitor that shows promising efficacy in HCC therapy. ABT-263 inhibits the anti-apoptotic activity of Bcl-2 and Bcl-xL, but not Mcl-1. Previous reports have shown that ABT-263 upregulates Mcl-1 in various cancer cells, which contributes to ABT-263 resistance in cancer therapy. However, the associated mechanisms are not well known. METHODS: Western blot, RNAi and CCK-8 assays were used to investigate the relationship between Mcl-1 upregulation and ABT-263 sensitivity in HCC cells. Real-time PCR and Western blot were used to detect Mcl-1 mRNA and protein levels. Luciferase reporter assay and RNA synthesis inhibition assay were adopted to analyze the mechanism of Mcl-1 mRNA upregulation. Western blot and the inhibition assays for protein synthesis and proteasome were used to explore the mechanisms of ABT-263-enhanced Mcl-1 protein stability. Trypan blue exclusion assay and flow cytometry were used to examine cell death and apoptosis. RESULTS: ABT-263 upregulated Mcl-1 mRNA and protein levels in HCC cells, which contributes to ABT-263 resistance. ABT-263 increased the mRNA level of Mcl-1 in HCC cells by enhancing the mRNA stability without influencing its transcription. Furthermore, ABT-263 increased the protein stability of Mcl-1 through promoting ERK- and JNK-induced phosphorylation of Mcl-1Thr163 and increasing the Akt-mediated inactivation of GSK-3ß. Additionally, the inhibitors of ERK, JNK or Akt sensitized ABT-263-induced apoptosis in HCC cells. CONCLUSIONS: ABT-263 increases Mcl-1 stability at both mRNA and protein levels in HCC cells. Inhibition of ERK, JNK or Akt activity sensitizes ABT-263-induced apoptosis. This study may provide novel insights into the Bcl-2-targeted cancer therapeutics.

CircRNA regulates the liquid-liquid phase separation of ATG4B, a novel strategy to inhibit cancer metastasis?

Anoikis is a common programmed death for most of detached cells, but cancer cells can obtain anoikis resistance to facilitate their distant metastasis through the circulation system. Researches have indicated that enhanced autophagic flux accounts for the survival of many cancer cells under detached conditions. Targeting ATG4B, the key factor of autophagy progress, can inhibit cancer metastasis in vitro, but ATG4B-deficient mice are susceptible to many serious diseases, which indicates the potential uncontrolled side effects of direct targeting of ATG4B. In our recent research, we confirmed that ATG4B is a novel RNA binding protein in the gastric cancer (GC) cell. It interacts with circSPECC1 which consequently facilitates the liquid-liquid phase separation and ubiquitination of ATG4B. Additionally, the m6A reader ELAVL1 inhibits the expression of circSPECC1 to enhance the expression of ATG4B and anoikis resistance of GC cells. Further, we screened out an FDA-approved compound, lopinavir, to restore circSPECC1 abundance and suppress GC metastasis. In conclusion, our research identified a novel signal pathway (ELAVL1-circSPECC1-ATG4B-autophagy) to facilitate anoikis resistance and metastasis of GC cells and screened out a compound with clinical application potential to block this pathway, providing a novel strategy for the prevention of GC metastasis.

Inhibition of KIAA1429/HK1 axis enhances the sensitivity of liver cancer cells to sorafenib by regulating the Warburg effect.

N6-methyladenosine (m6A) serves as the most abundant posttranscription modification. However, the role of m6A in tumorigenesis and chemotherapeutic drugs sensitivity remains largely unclear. Present research focuses on the potential function of the m6A writer KIAA1429 in tumor development and sorafenib sensitivity in liver cancer. We found that the level of KIAA1429 was significantly elevated in liver cancer tissues and cells and was closely associated with poorer prognosis. Functionally, KIAA1429 promoted the proliferation and Warburg effect of liver cancer cells in vitro and in vivo. RNA-seq and MeRIP-seq analysis revealed the glycolysis was one of the most affected pathways by KIAA1429, and m6A-modified HK1 was the most likely targeted gene to regulate the Warburg effect. KIAA1429 depletion decreased Warburg effect and increased sorafenib sensitivity in liver cancer. Mechanistically, KIAA1429 could affect the m6A level of HK1 mRNA through directly binding with it. Moreover, KIAA1429 cooperated with the m6A reader HuR to enhance HK1 mRNA stability, thereby upregulating its expression. These findings demonstrated that KIAA1429/HK1 axis decreases the sensitivity of liver cancer cells to sorafenib by regulating the Warburg effect, which may provide a novel therapeutic target for liver cancer treatment.

KIAA1429 facilitates metastasis via m6A-YTHDC1-dependent RND3 down-regulation in hepatocellular carcinoma cells.

N6-methyladenosine (m6A), a dynamically reversible modification in eukaryotic RNAs, modulates gene expression and pathological processes in various tumors. KIAA1429, the largest component of the m6A methyltransferase complex, plays an important role in m6A modification. However, the underlying mechanism of KIAA1429 in hepatocellular carcinoma (HCC) remains largely unknown. Immunohistochemical assay was performed to examine the expression of KIAA1429 in HCC tissues. Transwell, wound healing and animal experiments were used to investigate the influence of KIAA1429 on cell migration and invasion. The mRNA high-throughput sequencing (RNA-seq) and methylated RNA immunoprecipitation sequencing (MeRIP-seq) were performed to screen the downstream target of KIAA1429. RNA stability assays, RNA immunoprecipitation assay (RIP), MeRIP-qPCR and luciferase assay were used to evaluate the relationship between KIAA1429 and the m6A-modified genes. Results showed that the expression level of KIAA1429 was significantly higher in HCC tissues than in adjacent tissues, and the upregulation of KIAA1429 could promote HCC metastasis in vitro and in vivo. Mechanistically, we confirmed that KIAA1429 negatively regulated the tumor suppressor, Rho family GTPase 3 (RND3), by decreasing its mRNA stability in coordination with the m6A reader YTHDC1. Moreover, we demonstrated that KIAA1429 could regulate the m6A modification of RND3 mRNA via its RNA binding domain. Our data indicated that KIAA1429 exerted its oncogenic role by inhibiting RND3 expression in an m6A-dependent manner, suggesting that KIAA1429 might be a potential prognostic biomarker and therapeutic target in HCC.

Oxyberberine sensitizes liver cancer cells to sorafenib via inhibiting NOTCH1-USP7-c-Myc pathway.

BACKGROUND: Sorafenib is the first-line therapy for patients with advanced-stage HCC, but its clinical cure rate is unsatisfactory due to adverse reactions and drug resistance. Novel alternative strategies to overcome sorafenib resistance are urgently needed. Oxyberberine (OBB), a major metabolite of berberine in vivo, exhibits potential antitumor potency in various human malignancies, including liver cancer. However, it remains unknown whether and how OBB sensitizes liver cancer cells to sorafenib. METHODS: Cell viability, trypan blue staining and flow cytometry assays were employed to determine the synergistic effect of OBB and sorafenib on killing HCC cells. PCR, western blot, co-immunoprecipitation and RNA interference assays were used to decipher the mechanism by which OBB sensitizes sorafenib. HCC xenograft models and clinical HCC samples were utilized to consolidate our findings. RESULTS: We found for the first time that OBB sensitized liver cancer cells to sorafenib, enhancing its inhibitory effect on cell growth and induction of apoptosis in vitro. Interestingly, we observed that OBB enhanced the sensitivity of HCC cells to sorafenib by reducing ubiquitin-specific peptidase 7 (USP7) expression, a well-known tumor-promoting gene. Mechanistically, OBB inhibited notch homolog 1-mediated USP7 transcription, leading to the downregulation of V-Myc avian myelocytomatosis viral oncogene homolog (c-Myc), which synergized with sorafenib to suppress liver cancer. Furthermore, animal results showed that cotreatment with OBB and sorafenib significantly inhibited the tumor growth of liver cancer xenografts in mice. CONCLUSIONS: These results indicate that OBB enhances the sensitivity of liver cancer cells to sorafenib through inhibiting notch homolog 1-USP7-c-Myc signaling pathway, which potentially provides a novel therapeutic strategy for liver cancer to improve the effectiveness of sorafenib.

Lopinavir enhances anoikis by remodeling autophagy in a circRNA-dependent manner.

Macroautophagy/autophagy-mediated anoikis resistance is crucial for tumor metastasis. As a key autophagy-related protein, ATG4B has been demonstrated to be a prospective anti-tumor target. However, the existing ATG4B inhibitors are still far from clinical application, especially for tumor metastasis. In this study, we identified a novel circRNA, circSPECC1, that interacted with ATG4B. CircSPECC1 facilitated liquid-liquid phase separation of ATG4B, which boosted the ubiquitination and degradation of ATG4B in gastric cancer (GC) cells. Thus, pharmacological addition of circSPECC1 may serve as an innovative approach to suppress autophagy by targeting ATG4B. Specifically, the circSPECC1 underwent significant m6A modification in GC cells and was subsequently recognized and suppressed by the m6A reader protein ELAVL1/HuR. The activation of the ELAVL1-circSPECC1-ATG4B pathway was demonstrated to mediate anoikis resistance in GC cells. Moreover, we also verified that the above pathway was closely related to metastasis in tissues from GC patients. Furthermore, we determined that the FDA-approved compound lopinavir efficiently enhanced anoikis and prevented metastasis by eliminating repression of ELAVL1 on circSPECC1. In summary, this study provides novel insights into ATG4B-mediated autophagy and introduces a viable clinical inhibitor of autophagy, which may be beneficial for the treatment of GC with metastasis.

A Novel Cargo Delivery System-AnCar-ExoLaIMTS Ameliorates Arthritis via Specifically Targeting Pro-Inflammatory Macrophages.

Macrophages are heterogenic phagocytic cells that play distinct roles in physiological and pathological processes. Targeting different types of macrophages has shown potent therapeutic effects in many diseases. Although many approaches are developed to target anti-inflammatory macrophages, there are few researches on targeting pro-inflammatory macrophages, which is partially attributed to their non-s pecificity phagocytosis of extracellular substances. In this study, a novel recombinant protein is constructed that can be anchored on an exosome membrane with the purpose of targeting pro-inflammatory macrophages via antigen recognition, which is named AnCar-ExoLaIMTS . The data indicate that the phagocytosis efficiencies of pro-inflammatory macrophages for different AnCar-ExoLaIMTS show obvious differences. The AnCar-ExoLaIMTS3 has the best targeting ability for pro-inflammatory macrophages in vitro and in vivo. Mechanically, AnCar-ExoLaIMTS3 can specifically recognize the leucine-rich repeat domain of the TLR4 receptor, and then enter into pro-inflammatory macrophages via the TLR4-mediated receptor endocytosis pathway. Moreover, AnCar-ExoLaIMTS3 can efficiently deliver therapeutic cargo to pro-inflammatory macrophages and inhibit the synovial inflammatory response via downregulation of HIF-1α level, thus ameliorating the severity of arthritis in vivo. Collectively, the work established a novel gene/drug delivery system that can specifically target pro-inflammatory macrophages, which may be beneficial for the treatments of arthritis and other inflammatory diseases.

Circular RNA vaccine in disease prevention and treatment.

CircRNAs are a class of single-stranded RNAs with covalently linked head-to-tail topology. In the decades since its initial discovery, their biogenesis, regulation, and function have rapidly disclosed, permitting a better understanding and adoption of them as new tools for medical applications. With the development of biotechnology and molecular medicine, artificial circRNAs have been engineered as a novel class of vaccines for disease treatment and prevention. Unlike the linear mRNA vaccine which applications were limited by its instability, inefficiency, and innate immunogenicity, circRNA vaccine which incorporate internal ribosome entry sites (IRESs) and open reading frame (ORF) provides an improved approach to RNA-based vaccination with safety, stability, simplicity of manufacture, and scalability. However, circRNA vaccines are at an early stage, and their optimization, delivery and applications require further development and evaluation. In this review, we comprehensively describe circRNA vaccine, including their history and superiority. We also summarize and discuss the current methodological research for circRNA vaccine preparation, including their design, synthesis, and purification. Finally, we highlight the delivery options of circRNA vaccine and its potential applications in diseases treatment and prevention. Considering their unique high stability, low immunogenicity, protein/peptide-coding capacity and special closed-loop construction, circRNA vaccine, and circRNA-based therapeutic platforms may have superior application prospects in a broad range of diseases.

Involvement of acetylation of ATG4B in controlling autophagy induction.

ATG4B, a cysteine protease promoting autophagosome formation by reversibly modifying Atg8-family proteins, plays a vital role in controlling macroautophagy/autophagy initiation in response to stress. However, the molecular mechanism underlying the regulation of ATG4B activity is far from well elucidated. In the current study, we firstly revealed that the acetylation level of ATG4B at lysine residue 39 (K39) is strongly involved in regulating its activity and autophagy. Specifically, SIRT2 deacetylates ATG4B K39, enhancing ATG4B activity and autophagic flux, which can be antagonized by EP300/p300. Starvation treatment contributes to EP300 suppression and SIRT2 activation, promoting the deacetylation of ATG4B K39, which leads to the elevation of ATG4B activity and finally autophagy initiation. Mechanistic investigation showed that starvation reduces CCNE (cyclin E), resulting in the downregulation of the CCNE-CDK2 protein complex, decreasing the phosphorylation of SIRT2 Ser331 and finally activating SIRT2. In addition, we confirmed that SIRT2 promotes autophagy via suppressing acetylation of ATG4B at K39 using sirt2 gene knockout (sirt2-/-) mice. Collectively, our results have revealed the acetylation-mediated regulation of ATG4B cysteine protease activity in autophagy initiation in response to nutritional deficiency.

Autophagy in the pathogenesis and therapeutic potential of post-traumatic osteoarthritis.

Autophagy, as a fundamental mechanism for cellular homeostasis, is generally involved in the occurrence and progression of various diseases. Osteoarthritis (OA) is the most common musculoskeletal disease that often leads to pain, disability and economic loss in patients. Post-traumatic OA (PTOA) is a subtype of OA, accounting for >12% of the overall burden of OA. PTOA is often caused by joint injuries including anterior cruciate ligament rupture, meniscus tear and intra-articular fracture. Although a variety of methods have been developed to treat acute joint injury, the current measures have limited success in effectively reducing the incidence and delaying the progression of PTOA. Therefore, the pathogenesis and intervention strategy of PTOA need further study. In the past decade, the roles and mechanisms of autophagy in PTOA have aroused great interest in the field. It was revealed that autophagy could maintain the homeostasis of chondrocytes, reduce joint inflammatory level, prevent chondrocyte death and matrix degradation, which accordingly improved joint symptoms and delayed the progression of PTOA. Moreover, many strategies that target PTOA have been revealed to promote autophagy. In this review,  we summarize the roles and mechanisms of autophagy in PTOA and the current strategies for PTOA treatment that depend on autophagy regulation, which may be beneficial for PTOA patients in the future.

Dihydroergotamine mesylate enhances the anti-tumor effect of sorafenib in liver cancer cells.

Liver cancer is the most common and frequentlyoccurring cancer. In addition to radiotherapy, chemotherapy and surgery are recommended as part of liver cancer treatment. The efficacy of sorafenib and sorafenib-based combination treatment against tumors has been verified. Although, clinical trials have revealed that some individuals are not sensitive to sorafenib therapy, and current therapeutic approaches are ineffective. Consequently, it is urgent to explore effective drug combinations and innovative techniques for increasing the effectiveness of sorafenib in the curing of liver tumor. Herein, we show that dihydroergotamine mesylate (DHE), an anti-migraine agent, could effectively suppress liver cancer cells proliferation by inhibiting STAT3 activation. However, DHE can enhance the protein stability of Mcl-1 by activating ERK, making DHE less effective in apoptosis induction. Specifically, DHE enhances the effects of sorafenib on liver cancer cells, such as decreased viability and increased apoptosis. Furthermore, the mixture of sorafenib and DHE could enhance DHE-triggered STAT3 suppression and inhibit DHE-mediated ERK-Mcl-1 pathway activation. In vivo, the combination of sorafenib with DHE produced a substantial synergy in suppressing tumour growth and causing apoptosis, ERK inhibition and Mcl-1 degradation. These findings suggest that DHE can effectively inhibit cell proliferation and enhance sorafenib anti-cancer activity in liver cancer cells. The current study provides some new insights that DHE asa novel anti-liver cancer therapeutic agent has been shown to improve treatment outcomes of sorafenib, which might be helpful in order to advance sorafenib in liver cancer therapeutics.

Cir-DNA Sequencing Revealed the Landscape of Extrachromosomal Circular DNA in Articular Cartilage and the Potential Roles in Osteoarthritis.

OBJECTIVE: Extrachromosomal circular DNA (eccDNA) has been shown to be involved in several physiological and pathological processes including immunity, inflammation, aging, and tumor. However, the expression of eccDNA in cartilage has not been reported until now. In this study, we aimed to investigate the landscape of eccDNA in articular cartilage and analyze the potential roles in osteoarthritis (OA). METHODS: The samples of articular cartilage were obtained from total knee arthroplasty (TKA) donors with OA. The mitochondrial DNA (mtDNAs) and the linear DNAs from chondrocytes of articular cartilage were removed. Then the eccDNAs were enriched for cir-DNA sequencing. After quality control evaluation, we systematically revealed the identified eccDNA data including size distribution, the size range, and sequence pattern. Moreover, we explored and discussed the potential roles of eccDNA in OA via motif analysis and Gene Ontology (GO)/Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. RESULTS: The chondrocytes from OA cartilage contained an abundance of eccDNAs, which was termed as OC-eccDNAs (OA cartilage-derived eccDNA). The characteristics of OC-eccDNAs were tissue-specific, including the distribution, the size range, and sequence pattern. Moreover, the functional analysis indicated that eccDNA may be involved in the homeostasis maintenance of chondrocytes and participated in the process of OA. CONCLUSIONS: Our data first showed the landscape of eccDNA in articular cartilage and preliminarily indicated the potential roles of eccDNA in OA.

(-)-Gossypol enhances the anticancer activity of epirubicin via downregulating survivin in hepatocellular carcinoma.

Epirubicin (EPI)-based transarterial chemoembolization is an effective therapy for advanced hepatocellular carcinoma (HCC). However, EPI-induced survivin expression limits its tumor-killing potential in HCC. Interestingly, (-)-gossypol ((-)-Gsp), a male contraceptive, suppresses various malignancies. More importantly, (-)-Gsp also holds promise for enhancing the antitumor effects of chemotherapy in numerous cancer types. In the present study, we demonstrated for the first time that (-)-Gsp-sensitized EPI inhibited cell growth and induced apoptosis of HCC cells in vitro. Furthermore, (-)-Gsp sensitized EPI by attenuating the EPI-elevated survivin protein levels. Mechanistic studies showed that EPI stimulated survivin protein synthesis by promoting translation initiation, which was alleviated by (-)-Gsp mainly through suppressing the AKT-4EBP1/p70S6K-survivin and ERK-4EBP1-survivin pathways. HCC xenograft experiments in nude mice also showed that (-)-Gsp treatment acted synergistically with EPI to repress xenograft tumor growth. Overall, our proof-of-concept results may pave the way for novel strategies for the treatment of HCC based on the combination of EPI and (-)-Gsp.

Deacetylation of ATG4B promotes autophagy initiation under starvation.

Eukaryotes initiate autophagy when facing environmental changes such as a lack of external nutrients. However, the mechanisms of autophagy initiation are still not fully elucidated. Here, we showed that deacetylation of ATG4B plays a key role in starvation-induced autophagy initiation. Specifically, we demonstrated that ATG4B is activated during starvation through deacetylation at K39 by the deacetylase SIRT2. Moreover, starvation triggers SIRT2 dephosphorylation and activation in a cyclin E/CDK2 suppression-dependent manner. Meanwhile, starvation down-regulates p300, leading to a decrease in ATG4B acetylation at K39. K39 deacetylation also enhances the interaction of ATG4B with pro-LC3, which promotes LC3-II formation. Furthermore, an in vivo experiment using Sirt2 knockout mice also confirmed that SIRT2-mediated ATG4B deacetylation at K39 promotes starvation-induced autophagy initiation. In summary, this study reveals an acetylation-dependent regulatory mechanism that controls the role of ATG4B in autophagy initiation in response to nutritional deficiency.