Hepatic IRE1α-XBP1 signaling promotes GDF15-mediated anorexia and body weight loss in chemotherapy.

Platinum-based chemotherapy drugs can lead to the development of anorexia, a detrimental effect on the overall health of cancer patients. However, managing chemotherapy-induced anorexia and subsequent weight loss remains challenging due to limited effective therapeutic strategies. Growth differentiation factor 15 (GDF15) has recently gained significant attention in the context of chemotherapy-induced anorexia. Here, we report that hepatic GDF15 plays a crucial role in regulating body weight in response to chemo drugs cisplatin and doxorubicin. Cisplatin and doxorubicin treatments induce hepatic Gdf15 expression and elevate circulating GDF15 levels, leading to hunger suppression and subsequent weight loss. Mechanistically, selective activation by chemotherapy of hepatic IRE1α-XBP1 pathway of the unfolded protein response (UPR) upregulates Gdf15 expression. Genetic and pharmacological inactivation of IRE1α is sufficient to ameliorate chemotherapy-induced anorexia and body weight loss. These results identify hepatic IRE1α as a molecular driver of GDF15-mediated anorexia and suggest that blocking IRE1α RNase activity offers a therapeutic strategy to alleviate the adverse anorexia effects in chemotherapy.

Promotion of healthy adipose tissue remodeling ameliorates muscle inflammation in a mouse model of sarcopenic obesity.

A large subset of elders is classified as having sarcopenic obesity, a prevalence of obesity in combination with sarcopenia which places an aging population at the risk of adverse health consequences from both conditions. However, its complex etiology has restrained the development of effective therapeutic strategies. Recent progress has highlighted that the mode by which adipose tissue (AT) remodels is a determinant of metabolic health in the context of obesity. Healthy AT remodeling confers metabolic protection including insulin-sensitizing and anti-inflammatory effects to non-adipose tissues including skeletal muscle. Here, we employed a doxycycline-inducible adipocyte Hif1a knockout system to evaluate the muscle-protective effects associated with HIF1α inactivation-induced healthy AT remodeling in a model of sarcopenic obesity. We found that adipocyte HIF1α inactivation leads to improved AT metabolic health, reduced serum levels of lipids and pro-inflammatory cytokines, and increase of circulating adipokine (APN) in ovariectomized obese mice fed with obesogenic high-fat diet (HFD). Concomitantly, muscle inflammation is evidently lower in obese OVX mice when adipocyte HIF1α is inactivated. Furthermore, these protective effects against muscle inflammation can be mimicked by the administration of adiponectin receptor agonist AdipoRon. Collectively, our findings underscore the importance of AT metabolic health in the context of concurrent sarcopenia and obesity, and promotion of healthy AT remodeling may represent a new therapeutic strategy to improve muscle health in sarcopenic obesity.

LncRNA-HOTAIR activates autophagy and promotes the imatinib resistance of gastrointestinal stromal tumor cells through a mechanism involving the miR-130a/ATG2B pathway.

Gastrointestinal stromal tumors (GISTs) are common neoplasms of the gastrointestinal tract that can be treated successfully using C-kit target therapy and surgery; however, imatinib chemoresistance is a major barrier to success in therapy. The present study aimed to discover alternative pathways in imatinib-resistant GISTs. Long noncoding RNAs (lncRNAs) are newly discovered regulators of chemoresistance. Previously, we showed that the lncRNA HOTAIR was upregulated in recurrent GISTs. In this study, we analyzed differentially expressed lncRNAs after imatinib treatment and found that HOTAIR displayed the largest increase. The distribution of HOTAIR in GISTs was shifted from nucleus to cytoplasm after imatinib treatments. The expression of HOTAIR was validated as related to drug sensitivity through Cell Counting Kit-8 assays. Moreover, HOTAIR was associated strongly with cell autophagy and regulated drug sensitivity via autophagy. Mechanistically, HOTAIR correlated negatively with miRNA-130a in GISTs. The downregulation of miRNA-130a reversed HOTAIR-small interfering RNA-induced suppression of autophagy and imatinib sensitivity. We identified autophagy-related protein 2 homolog B (ATG2B) as a downstream target of miR-130a and HOTAIR. ATG2B downregulation reversed the effect of pEX-3-HOTAIR/miR-130a inhibitor on imatinib sensitivity. Finally, HOTAIR was shown to influence the autophagy and imatinib sensitivity of GIST cells in mouse tumor models. Our results suggested that HOTAIR targets the ATG2B inhibitor miR-130a to upregulate the level of cell autophagy so that promotes the imatinib resistance in GISTs.

Knockdown of eukaryotic translation initiation factor 5A2 enhances the therapeutic efficiency of doxorubicin in hepatocellular carcinoma cells by triggering lethal autophagy.

Hepatocellular carcinoma (HCC) is an invasive malignant neoplasm with a poor prognosis. The development of chemoresistance severely obstructs the chemotherapeutic efficiency of HCC treatment. Therefore, understanding the mechanisms of chemoresistance is important for improving the outcomes of patients with HCC. Eukaryotic translation initiation factor 5A2 (eIF5A2), which is considered to be an oncogene, has been reported to mediate chemoresistance in various types of cancer; however, its precise role in HCC remains unclear. Accumulating evidence has suggested that autophagy serves a dual role in cancer chemotherapy. The present study aimed to investigate the role of autophagy in eIF5A2­mediated doxorubicin resistance in HCC. High expression levels of eIF5A2 in human HCC tissues were observed by immunohistochemistry using a tissue microarray, which was consistent with the results of reverse transcription­quantitative PCR analysis in paired HCC and adjacent healthy tissues. HCC patient­derived tumor xenograft mouse model was used for the in vivo study, and knockdown of eIF5A2 effectively enhanced the efficacy of doxorubicin chemotherapy compared with that in the control group. Notably, eIF5A2 served as a repressor in regulating autophagy under chemotherapy. Silencing of eIF5A2 induced doxorubicin sensitivity in HCC cells by triggering lethal autophagy. In addition, 5­ethynyl­2'­deoxyuridine, lactate dehydrogenase release assay and calcein­AM/PI staining were used to determine the enhanced autophagic cell death induced by the silencing of eIF5A2 under doxorubicin treatment. Suppression of autophagy attenuated the sensitivity of HCC cells to doxorubicin induced by eIF5A2 silencing. The results also demonstrated that knockdown of the Beclin 1 gene, which is an autophagy regulator, reversed the enhanced autophagic cell death and doxorubicin sensitivity induced by eIF5A2 silencing. Taken together, these results suggested eIF5A2 may mediate the chemoresistance of HCC cells by suppressing autophagic cell death under chemotherapy through a Beclin 1­dependent pathway, and that eIF5A2 may be a novel potential therapeutic target for HCC treatment.

Chimeric peptide supramolecular nanoparticles for plectin-1 targeted miRNA-9 delivery in pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal disease with poor prognosis. Insights into the roles of MicroRNAs (miRNAs) in diseases, particularly in cancer, have made miRNAs attractive tools and targets for novel therapeutic approaches. Methods: Here, we employed a novel chimeric peptide supramolecular nanoparticle delivery system for plectin-1 (PL-1)-targeted PDAC-specific miR-9 delivery in vitro and in pancreatic cancer patient-derived xenograft (PDX) model. RT-PCR and immunohistochemistry (IHC) were conducted to detect the expression pattern of eIF5A2. mRFP-GFP-LC3 fluorescence microscopy and Western blot were carried out to determine autophagy. Luciferase reporter assays were performed to elucidate the regulatory role of miR-9/eIF5A2 axis. Results: PL-1/miR-9 nanocomplexes dramatically improve the anticancer effect of doxorubicin through downregulating eIF5A2 expression to inhibit autophagy and induce apoptosis in PDAC therapy in vivo. Mechanistically, miR-9 directly targets the eIF5A2 transcript by binding to its 3'-untranslated region (3'-UTR) to reduce the expression levels and the secreted protein of eIF5A2 in PDAC cells. Conclusion: PL-1/miR-9 nanoparticles can be used as a novel promising anti-cancer strategy with tumor targeting and miR-9/eIF5A2 may serve as a new potential therapeutic target for future synergic therapy against human PDAC.

miR-223 overexpression inhibits doxorubicin-induced autophagy by targeting FOXO3a and reverses chemoresistance in hepatocellular carcinoma cells.

Doxorubicin is conventionally used in chemotherapy against hepatocellular carcinoma (HCC), but acquired resistance developed during long-term therapy limits its benefits. Autophagy, a conserved catabolic process for cellular self-protection and adaptation to the changing environment, is regarded as a potential clinical target to overcome doxorubicin resistance. In this study, the potential role of miR-223 in modulating doxorubicin-induced autophagy and sensitivity were evaluated in four transfected human HCC cell lines, and the in vivo relevance was assessed using a mouse xenograft model of HCC. We found that the well-defined miR-223 is expressed at low levels in doxorubicin treated HCC cells and that miR-223 overexpression inhibits the doxorubicin-induced autophagy that contributes to chemoresistance. Blockade of autophagic flux by chloroquine resulted in the failure of miR-223 inhibitor to suppress doxorubicin sensitivity of HCC cells. We further identified FOXO3a as a direct downstream target of miR-223 and primary mediator of the regulatory effect of miR-223 on doxorubicin-induced autophagy and chemoresistance in HCC cells. Finally, we confirmed the enhancement of doxorubicin sensitivity by agomiR-223 in xenograft models of HCC. These findings establish a novel miRNA-based approach for autophagy interference to reverse doxorubicin resistance in future chemotherapy regimens against human HCC.

CRISPR/Cas9-Mediated Treatment Ameliorates the Phenotype of the Epidermolytic Palmoplantar Keratoderma-like Mouse.

CRISPR/Cas9 has been confirmed as a distinctly efficient, simple-to-configure, highly specific genome-editing tool that has been used to treat monogenetic disorders. Epidermolytic palmoplantar keratoderma (EPPK) is a common autosomal dominant keratin disease resulting from dominant-negative mutation of the KRT9 gene, and it has no effective therapy. We performed CRISPR/Cas9-mediated treatment on a knockin (KI) transgenic mouse model that carried a small indel heterozygous mutation of Krt9, c.434delAinsGGCT (p.Tyr144delinsTrpLeu), which caused a humanized EPPK-like phenotype. The mutation within exon 1 of Krt9 generated a novel protospacer adjacent motif site, TGG, for Cas9 recognition and cutting. By delivering lentivirus vectors (LVs) encoding single-guide RNAs (sgRNAs) and Cas9 that targeted Krt9 sequence into HeLa cells engineered to constitutively express wild-type and mutant keratin 9 (K9), we found the sgRNA was highly effective in reducing expression of the mutant K9 protein in vitro. We injected the LV into the fore-paws of adult KI-Krt9 mice three times every 8 days and found that the expression of K9 decreased ∼14.6%. The phenotypic mitigation was revealed by restoration of the abnormal differentiation and aberrant proliferation of the epidermis. Our data are the first to show that CRISPR/Cas9 is a potentially powerful therapeutic option for EPPK and other PPK subtypes.

Eukaryotic translation initiation factor 5A2 regulates the migration and invasion of hepatocellular carcinoma cells via pathways involving reactive oxygen species.

Eukaryotic translation initiation factor 5A2 (eIF5A2) has been identified as a critical gene in tumor metastasis. Research has suggested that reactive oxygen species (ROS) serve as signaling molecules in cancer cell proliferation and migration. However, the mechanisms linking eIF5A2 and ROS are not fully understood. Here, we investigated the effects of ROS on the eIF5A2-induced epithelial-mesenchymal transition (EMT) and migration in six hepatocellular carcinoma (HCC) cell lines. Western hybridization, siRNA transfection, transwell migration assays, wound-healing assays, and immunofluorescence analysis were used. The protein levels of eIF5A2 in tumor and adjacent tissue samples from 90 HCC patients with detailed clinical, pathological, and clinical follow-up data were evaluated. Overexpression of eIF5A2 was found in cancerous tissues compared with adjacent tissues. We found that eIF5A2 overexpression in HCC was associated with reduced overall survival. Knockdown of eIF5A2 and intracellular reduction of ROS significantly suppressed the invasion and metastasis of HCC cells. Interestingly, N1-guanyl-1, 7-diaminoheptane (GC7) suppressed the intracellular ROS levels. After blocking the EMT, administration of GC7 or N-acetyl-L-cysteine did not reduce cell migration further. Based on the experimental data, we concluded that inhibition of eIF5A2 alters progression of the EMT to decrease the invasion and metastasis of HCC cells via ROS-related pathways.

A Small Indel Mutant Mouse Model of Epidermolytic Palmoplantar Keratoderma and Its Application to Mutant-specific shRNA Therapy.

Epidermolytic palmoplantar keratoderma (EPPK) is a relatively common autosomal-dominant skin disorder caused by mutations in the keratin 9 gene (KRT9), with few therapeutic options for the affected so far. Here, we report a knock-in transgenic mouse model that carried a small insertion-deletion (indel) mutant of Krt9, c.434delAinsGGCT (p.Tyr144delinsTrpLeu), corresponding to the human mutation KRT9/c.500delAinsGGCT (p.Tyr167delinsTrpLeu), which resulted in a human EPPK-like phenotype in the weight-stress areas of the fore- and hind-paws of both Krt9(+/mut) and Krt9(mut/mut) mice. The phenotype confirmed that EPPK is a dominant-negative condition, such that mice heterozygotic for the K9-mutant allele (Krt9(+/mut)) showed a clear EPPK-like phenotype. Then, we developed a mutant-specific short hairpin RNA (shRNA) therapy for EPPK mice. Mutant-specific shRNAs were systematically identified in vitro using a luciferase reporter gene assay and delivered into Krt9(+/mut) mice. shRNA-mediated knockdown of mutant protein resulted in almost normal morphology and functions of the skin, whereas the same shRNA had a negligible effect in wild-type K9 mice. Our results suggest that EPPK can be treated by gene therapy, and this has significant implications for future clinical application.