LINC00982-encoded protein PRDM16-DT regulates CHEK2 splicing to suppress colorectal cancer metastasis and chemoresistance.

Metastasis is one of the key factors of treatment failure in late-stage colorectal cancer (CRC). Metastatic CRC frequently develops resistance to chemotherapeutic agents. This study aimed to identify the novel regulators from "hidden" proteins encoded by long noncoding RNAs (lncRNAs) involved in tumor metastasis and chemoresistance. Methods: CRISPR/Cas9 library functional screening was employed to identify the critical suppressor of cancer metastasis in highly invasive CRC models. Western blotting, immunofluorescence staining, invasion, migration, wound healing, WST-1, colony formation, gain- and loss-of-function experiments, in vivo experimental metastasis models, multiplex immunohistochemical staining, immunohistochemistry, qRT-PCR, and RT-PCR were used to assess the functional and clinical significance of FOXP3, PRDM16-DT, HNRNPA2B1, and L-CHEK2. RNA-sequencing, co-immunoprecipitation, qRT-PCR, RT-PCR, RNA affinity purification, RNA immunoprecipitation, MeRIP-quantitative PCR, fluorescence in situ hybridization, chromatin immunoprecipitation and luciferase reporter assay were performed to gain mechanistic insights into the role of PRDM16-DT in cancer metastasis and chemoresistance. An oxaliplatin-resistant CRC cell line was established by in vivo selection. WST-1, colony formation, invasion, migration, Biacore technology, gain- and loss-of-function experiments and an in vivo experimental metastasis model were used to determine the function and mechanism of cimicifugoside H-1 in CRC. Results: The novel protein PRDM16-DT, encoded by LINC00982, was identified as a cancer metastasis and chemoresistance suppressor. The down-regulated level of PRDM16-DT was positively associated with malignant phenotypes and poor prognosis of CRC patients. Transcriptionally regulated by FOXP3, PRDM16-DT directly interacted with HNRNPA2B1 and competitively decreased HNRNPA2B1 binding to exon 9 of CHEK2, resulting in the formation of long CHEK2 (L-CHEK2), subsequently promoting E-cadherin secretion. PRDM16-DT-induced E-cadherin secretion inhibited fibroblast activation, which in turn suppressed CRC metastasis by decreasing MMP9 secretion. Cimicifugoside H-1, a natural compound, can bind to LEU89, HIS91, and LEU92 of FOXP3 and significantly upregulated PRDM16-DT expression to repress CRC metastasis and reverse oxaliplatin resistance. Conclusions: lncRNA LINC00982 can express a new protein PRDM16-DT to function as a novel regulator in cancer metastasis and drug resistance of CRC. Cimicifugoside H-1 can act on the upstream of the PRDM16-DT signaling pathway to alleviate cancer chemoresistance.

lncRNA MEG8 is downregulated in osteoarthritis and regulates chondrocyte cell proliferation, apoptosis and inflammation.

Long noncoding RNA (lncRNA) maternally expressed 8, small nucleolar RNA host gene (MEG8) has been widely reported for its pro-proliferative, anti-apoptotic and anti-inflammatory effects in diverse diseases. The aim of the present study was to investigate the effects and underlying mechanism of MEG8 on IL-1ß-stimulated human osteoarthritis (OA) chondrocytes. C28/I2 chondrocytes were cultured under the stimulation of IL-1ß to establish a cellular model of OA. Functional assays involving Cell Counting Kit-8 and flow cytometry were performed to determine proliferation and apoptosis in the cells. The protein expression levels of caspase-3 and inflammatory cytokines were detected using cell-based ELISA. The expression levels of PI3K/AKT pathway-related proteins were evaluated by western blotting. It was identified that MEG8 expression was increased in the cartilage of patients with OA and in IL-1ß-treated C28/I2 cells. In C28/I2 cells, silencing of MEG8 expression noticeably triggered IL-1ß-induced proliferation suppression, cell death and an inflammatory response. However, transfection with MEG8 displayed adverse effects. Furthermore, MEG8 overexpression prevented IL-1ß-induced activation of the PI3K/AKT signaling pathway in C28/I2 cells. These data demonstrated that MEG8 exerted protective effects against IL-1ß-induced apoptosis and inflammation of OA chondrocytes by regulating the PI3K/AKT signaling pathway. Thus, the present study demonstrates that MEG8 might be a promising target for the treatment of OA.

Knockdown of FBXO39 inhibits proliferation and promotes apoptosis of human osteosarcoma U-2OS cells.

F-box proteins are essential components of the Skp-cullin-F-box complex (a type of E3 ubiquitin ligase), and participate in cell cycle and immune responses through the ubiquitin proteasome system. F-box protein 39 (FBXO39) belongs to the F-box family, which has been reported to be associated with cancer oncogenesis and progression. The present study aimed to investigate the role of FBXO39 in osteosarcoma (OS) cell proliferation and apoptosis in vitro. It was demonstrated that U-2OS cells exhibited high expression of FBXO39 compared with HOS and SaOS-2 osteosarcoma cells. Thus, knockdown of FBXO39 was performed using lentivirus-mediated short hairpin RNA (shRNA) transfection to validate the effect of FBXO39 in U-2OS cells. Western blotting and RT-qPCR analysis were used to confirm the efficiency of infection by analyzing the expression level of FBXO39. Using Celigo-based cell counting and MTT assays, it was demonstrated that FBXO39 knockdown significantly reduced the rate of cell proliferation compared with control. Caspase 3/7 activity assays and fluorescence-activated cell sorting confirmed the induction of apoptosis in U-2OS cells following FBXO39 knockdown. In conclusion, it was demonstrated that FBXO39 knockdown may significantly inhibit proliferation and promote apoptosis of U-2OS cells. Thus, FBXO39 may serve an important role in OS progression.

Clinicopathological and prognostic significance of PD-L1 expression in sarcoma: A systematic review and meta-analysis.

Programmed cell death-ligands 1 (PD-L1) is a key immune checkpoint protein and a promising therapeutic target for malignancy tumor immunotherapy. The prognostic value of PD-L1 in patients with bone and soft tissue sarcoma remains controversial. Therefore, this meta-analysis is conducted to evaluate the associations of PD-L1 expression with overall survival (OS), progression-free survival (PFS), and clinicopathological characteristics of sarcomaA comprehensive literature search of PubMed, Web of Science, Embase, and Cochrane Library was conducted for relevant studies. A total of 14 studies published from 2013 to 2017 were included. The pooled hazard ratios (HRs) with 95% confidence intervals (CIs) were extracted from included studies to assess the association between PD-L1 expression and OS, PFS of patients with sarcoma. Other relevant data were extracted to evaluate the correlations of PD-L1 expression with risk and clinicopathological characteristics of sarcoma. Stata 12.0 software was applied to calculate the strength of association between PD-L1 expression and sarcoma.In total, 14 articles containing 15 independent studies and 1,451 patients were included in this meta-analysis. We found that the high PD-L1 expression was associated with poorer overall survival (HR 1.27, 95% CI: 0.70-1.84 P = .000) and poorer events-free survival (HR 2.05, 95% CI: 1.55-2.70, P = .000) in bone and soft-tissue sarcoma patients. Additionally, we conducted subgroup analysis according to histology type, ethnicity, target of PD-L1 assessment, cutoff, the significant correlations with poor overall survival and events-free survival were also observed. In contrast none of the clinicopathological characteristics (gender, age, tumor site, tumor grade, tumor depth, tumor necrosis rate, metastasis, recurrence, chemotherapy, radiotherapy) was found to be associated with PD-L1 expression in our analysis.The findings from this meta-analysis indicate that PD-L1 expression might be a useful predicative factor of poor prognosis for patients with bone and soft tissue sarcoma.