MIIP inhibits the growth of prostate cancer via interaction with PP1α and negative modulation of AKT signaling.

BACKGROUND: Over-activation of phosphatidylinositol 3-kinase (PI3K)-AKT-mammalian target of rapamycin (mTOR) signaling pathway is one of important mechanisms to promote castration resistant prostate cancer, the final stage of prostate cancer (PCa). Dysregulation of PP1-meditaed AKT dephosphorylation might contribute to such an event but is not fully understood. As a newly identified tumor suppressor, MIIP exerts its role in various types of cancer but has not been investigated in PCa. RESULTS: We first demonstrated that overexpression of migration and invasion inhibitory protein (MIIP) in human PCa cell lines suppresses their growth while knockdown of MIIP does the opposite in vitro. Although MIIP has no effect on the expression of AR and its target genes or the nuclear translocation of AR in AR-positive PCa cells, MIIP overexpression significantly inhibits activation of AKT-mTOR pathway in both AR- positive and negative PCa cells whereas knockdown of MIIP enhances AKT-mTOR signaling. Using Western blot, immunofluorescence co-localization and co-immunoprecipitation analysis, we found that MIIP interacts with PP1α via its C-terminal part but does not affect its protein level. Importantly, silence of PP1α reversed the inhibitory effect of MIIP on AKT phosphorylation and cell growth in PCa cell lines, while MIIP∆C, which is incapable of interacting with PP1α, loses MIIP's effect, suggesting that MIIP exerts its roles via interaction with PP1α. Further, MIIP overexpression inhibits the growth of both AR- positive and negative PCa xenograft in nude mice. Finally, immunohistochemical staining of PCa tissue microarray showed that MIIP expression level is downregulated in PCa and negatively correlated with Gleason score of PCa. CONCLUSION: We discovered that MIIP is a novel suppressor of oncogenic AKT-mTOR signaling in PCa by facilitating PP1-meditaed AKT dephosphorylation. Our study further emphasized the tumor suppressive role of MIIP and illustrated a novel mechanism.

Correction to: MIIP inhibits the growth of prostate cancer via interaction with PP1α and negative modulation of AKT signaling.

Following publication of the original article [1], the authors reported that the given name of Qinhao Wang was incorrectly published as Qinghao Wang. The original article has been corrected.

GOLM1 promotes prostate cancer progression through activating PI3K-AKT-mTOR signaling.

BACKGROUND: Prostate cancer (PCa) is the most commonly diagnosed cancer in men. Various molecular mechanisms account for PCa progression and elucidation of these mechanisms is key for selection of optimal therapies and improvement of patient outcome. Golgi membrane protein 1 (GOLM1) has been identified as a novel biomarker for PCa, but its biological functions and molecular mechanisms remain poorly understood. METHOD: GOLM1 expression was determined in PCa by tissue microarrays (TMAs) and real-time RT-PCR, Western blot, and immunohistochemistry (IHC) analyses. To investigate GOLM1 functions in vitro and in vivo, we overexpressed and knocked down GOLM1 in PCa cell lines and established xenograft mice models. A series of cytological function assays were used to determine the role of GOLM1 in cell proliferation, migration, invasion, and apoptosis. PI3K-AKT-mTOR signaling pathway downstream of GOLM1 was detected by Western blot and IHC analyses. RESULT: GOLM1 expression is up-regulated in PCa of all stages and grades. GOLM1 promotes proliferation, migration and invasion, and inhibits apoptosis in PCa cell lines (DU145, PC3, and CWR22Rv1) and xenograft mice models. Moreover, PI3K-AKT-mTOR signaling is positively regulated by GOLM1, whereas PI3 K inhibitor BKM120 significantly abrogates the oncogenic functions of GOLM1. CONCLUSION: GOLM1 acts as a critical oncogene by promoting PCa cell proliferation, migration and invasion, and inhibiting apoptosis. GOLM1 plays oncogenic functions mainly through activating PI3K-AKT-mTOR signaling pathway. Therefore, agents that block PI3K-AKT-mTOR signaling pathway could be used in PCa patients with GOLM1 up-regulation.

NDRG2 overexpression suppresses hepatoma cells survival during metabolic stress through disturbing the activation of fatty acid oxidation.

Because of the high nutrient consumption and inadequate vascularization, solid tumor constantly undergoes metabolic stress during tumor development. Oncogenes and tumor suppressor genes participated in cancer cells' metabolic reprogramming. N-Myc downstream regulated gene 2 (NDRG2) is a recently identified tumor suppressor gene, but its function in cancer metabolism, particularly during metabolic stress, remains unclear. In this study, we found that NDRG2 overexpression significantly reduced hepatoma cell proliferation and enhanced cell apoptosis under glucose limitation. Moreover, NDRG2 overexpression aggravated energy imbalance and oxidative stress by decreasing the intracellular ATP and NADPH generation and increasing ROS levels. Strikingly, NDRG2 inhibited the activation of fatty acid oxidation (FAO), which preserves ATP and NADPH purveyance in the absence of glucose. Finally, mechanistic investigation showed that NDRG2 overexpression suppressed the glucose-deprivation induced AMPK/ACC pathway activation in hepatoma cells, whereas the expression of a constitutively active form of AMPK abrogated glucose-deprivation induced AMPK activation and cell apoptosis. Thus, as a negative regulator of AMPK, NDRG2 disturbs the induction of FAO genes by glucose limitation, leading to dysregulation of ATP and NADPH, and thus reduces the tolerance of hepatoma cells to glucose limitation.

Functional and clinical evidence that TAZ is a candidate oncogene in hepatocellular carcinoma.

Transcriptional co-activator with PDZ-binding motif (TAZ) has been reported to be associated with carcinogenesis. However, the cellular function of TAZ in human hepatocellular carcinoma (HCC) remains elusive. In this study, an immunohistochemistry analysis revealed that the expression of TAZ in cancer tissue samples from 180 HCC patients was significantly higher than that in adjacent normal tissues. In addition, TAZ overexpression was significantly correlated with aggressive tumor characteristics such as tumor size, TNM stage, lymph node or distant metastasis, histological differentiation, and recurrent HCC (P < 0.05). The Kaplan-Meier test showed that TAZ-positive expression was related to a poor prognosis compared to TAZ-negative expression (P < 0.05). Furthermore, the expression level of TAZ was generally correlated with the invasiveness of cancer cells. The overexpression of TAZ in the Huh7 cell line, which endogenously expresses TAZ at low levels, significantly promoted cell proliferation, migration and invasion and inhibited apoptosis, whereas RNA interference-mediated knockdown of TAZ in the highly invasive cell line MHCC-97H significantly suppressed cell proliferation, migration and invasion in vitro and tumor formation in vivo.

MIIP downregulation drives colorectal cancer progression through inducing peri-cancerous adipose tissue browning.

BACKGROUND: The enrichment of peri-cancerous adipose tissue is a distinctive feature of colorectal cancer (CRC), accelerating disease progression and worsening prognosis. The communication between tumor cells and adjacent adipocytes plays a crucial role in CRC advancement. However, the precise regulatory mechanisms are largely unknown. This study aims to explore the mechanism of migration and invasion inhibitory protein (MIIP) downregulation in the remodeling of tumor cell-adipocyte communication and its role in promoting CRC. RESULTS: MIIP expression was found to be decreased in CRC tissues and closely associated with adjacent adipocyte browning. In an in vitro co-culture model, adipocytes treated with MIIP-downregulated tumor supernatant exhibited aggravated browning and lipolysis. This finding was further confirmed in subcutaneously allografted mice co-injected with adipocytes and MIIP-downregulated murine CRC cells. Mechanistically, MIIP interacted with the critical lipid mobilization factor AZGP1 and regulated AZGP1's glycosylation status by interfering with its association with STT3A. MIIP downregulation promoted N-glycosylation and over-secretion of AZGP1 in tumor cells. Subsequently, AZGP1 induced adipocyte browning and lipolysis through the cAMP-PKA pathway, releasing free fatty acids (FFAs) into the microenvironment. These FFAs served as the primary energy source, promoting CRC cell proliferation, invasion, and apoptosis resistance, accompanied by metabolic reprogramming. In a tumor-bearing mouse model, inhibition of ß-adrenergic receptor or FFA uptake, combined with oxaliplatin, significantly improved therapeutic efficacy in CRC with abnormal MIIP expression. CONCLUSIONS: Our data demonstrate that MIIP plays a regulatory role in the communication between CRC and neighboring adipose tissue by regulating AZGP1 N-glycosylation and secretion. MIIP reduction leads to AZGP1 oversecretion, resulting in adipose browning-induced CRC rapid progression and poor prognosis. Inhibition of ß-adrenergic receptor or FFA uptake, combined with oxaliplatin, may represent a promising therapeutic strategy for CRC with aberrant MIIP expression.

[Regulation of PD-L1 posttranslational modification and its application progress in tumor immunotherapy].

The immune checkpoint, programmed death 1 ligand-1/programmed death -1 (PD-L1/PD-1), is one of the most promising targets for tumor immunotherapy. Overexpressed PD-L1 on the surface of tumor cells could bind to PD-1 on the surface of T cells and inhibit the T cell activation, triggering tumor-immune-escape; therapeutic strategies targeting PD-1/PD-L1 restore the cytotoxic function of immune cells in the tumors by blocking PD-1/PD-L1 interaction. The PD-L1 undergoes multi-level regulations in tumor cells. Among them, post-translational modifications (PTMs) of PD-L1 mainly including glycosylation, phosphorylation, ubiquitination, acetylation and palmitoylation, have attracted much attention in recent years. These modifications directly affect the stability, cellular localization and function of PD-L1, and subsequently regulate the T cell activation and tumor immunity. Therefore, intervention with PTMs of PD-L1 may serve as a new approach for anti-tumor immunity-escape therapy.

miR-145-5p Modulates Gefitinib Resistance by Targeting NRAS and MEST in Non-Small Cell Lung Cancer.

OBJECTIVE: microRNAs may play essential roles in the development and drug resistance of non-small cell lung cancer (NSCLC). However, their functions and mechanisms are not fully understood. Our goal was to define the role of miR-145-5p in the gefitinib resistance of NSCLC. MATERIALS AND METHODS: An A549 gefitinib-resistant cell line and xenograft nude mice were used in this study. The expression of miR-145-5p and its targets, NRAS and MEST, were detected and measured by qPCR, Western blot, RNA-FISH, or immunofluorescence analysis. RESULTS: miR-145-5p was downregulated in gefitinib-resistant A549 cells (A549/Gef R). Overexpression of miR-145-5p enhanced the sensitivity to gefitinib and inhibited cell proliferation and invasion in A549/Gef R. miR-145-5p was also significantly reduced in LUAD and LUSC clinical samples and closely associated with a favorable prognosis, according to the UALCAN and TCGA databases. Moreover, NRAS and MEST were found to be downstream target genes of miR-145-5p and to function as oncogenes in NSCLC samples, and gefitinib resistance could be improved following the interference of these two molecules. CONCLUSION: miR-145-5p improves the sensitivity of acquired gefitinib-resistant cells to gefitinib via inhibiting NRAS and MEST expression. The miR 145-5p-NRAS/MEST axis in NSCLC provides insights for the development of a NRAS/MEST targeting therapeutic approach to overcome gefitinib resistance in NSCLC patients.

MIIP inhibits clear cell renal cell carcinoma proliferation and angiogenesis via negative modulation of the HIF-2α-CYR61 axis.

OBJECTIVE: In various cancers, migration and invasion inhibitory protein (MIIP) is expressed at low level and is involved in cancer pathogenesis. Herein, we sought to explore the function of MIIP in clear cell renal cell carcinoma (ccRCC). METHODS: CCK-8, colony formation, cell cycle, and endothelial cell tube formation assays were performed to evaluate the roles of MIIP in ccRCC proliferation and angiogenesis. To explore the underlying mechanism, we conducted RNA-sequencing, GSEA, qRT-PCR, Western blot, ELISA, cell transfection, coimmunoprecipitation, and ubiquitination assays in ccRCC cell lines. Furthermore, xenograft tumor growth in nude mice, and Ki-67 and CD31 staining in xenograft tissues were examined. Finally, the association of MIIP expression with clinical pathology and the expression status of HIF-2α and cysteine-rich 61 (CYR61) were further analyzed in human RCC tissues through Western blot and immunohistochemistry. RESULTS: Both in vitro and in vivo functional experiments indicated that forced expression of MIIP inhibited ccRCC proliferation and angiogenesis, whereas silencing MIIP either in normal HK-2 cells or in ccRCC cells had the opposite effect (P < 0.05). Mechanistically, CYR61 was identified as a gene significantly downregulated by MIIP overexpression, and was required for the suppressive role of MIIP in ccRCC. MIIP was found to promote HSP90 acetylation and thus impair its chaperone function toward HIF-2α. Consequently, RACK1 binds HIF-2α and causes its ubiquitination and proteasomal degradation, thus decreasing the transcription of its target, CYR61. Finally, analyses of clinical samples demonstrated that MIIP is significantly downregulated in cancer vs. normal tissues in RCC cases, and its expression is negatively associated with histological grade, metastasis, the prognosis of patients with RCC, and the expression of HIF-2α and CYR61 (P < 0.05). CONCLUSIONS: MIIP is a novel tumor suppressor in ccRCC via negative regulation of HIF-2α-CYR61 axis.

MIIP inhibits EMT and cell invasion in prostate cancer through miR-181a/b-5p-KLF17 axis.

Growing evidence have shown that the migration and invasion inhibitory protein (MIIP, also known as IIp45) functions as a tumor suppressor and its expression is downregulated in several types of cancer, yet the function of MIIP in prostate cancer (PCa) and the underlying mechanism of action remains largely unknown. Here we demonstrated that MIIP acts as a suppressor of PCa by inhibiting epithelial-mesenchymal transition (EMT) and cell invasion. Overexpressing MIIP repressed cellular invasion of PC3 and DU145 in vitro, accompanied by a decrease of EMT-inducing factors, and an increase of E-cadherin and KLF17. Moreover, a stable MIIP knockdown in PCa cells promoted the tumor growth or bone osteolytic lesions, when xenografted subcutaneously or via tibia injection. Mechanistically, MIIP represses two onco-miRNAs, miR-181a-5p and miR-181b-5p, thus removing the inhibitory effect of these two miRNAs on their target KLF17, which functions as a negative regulator of EMT by directly suppressing the transcription of SNAIL1/2 and TWIST. Finally, by examining the expression of MIIP, miR-181a/b-5p, KLF17, and E-cadherin in paired cancer samples v.s. adjacent normal tissues from a cohort of human prostate cancer patients, we demonstrated that downregulation of MIIP was well associated with downregulation of KLF17 and E-cadherin, but upregulation of miR-181a/b-5p. The positive correlation between MIIP and KLF17 was also confirmed via immunohistochemical staining of a PCa tissue microarray. Taken together, our findings reveal a novel function of MIIP as an EMT inhibitor in PCa and illustrate the underlying molecular mechanisms, providing new insights into the tumor-suppressor role of MIIP.

IDH2 compensates for IDH1 mutation to maintain cell survival under hypoxic conditions in IDH1­mutant tumor cells.

Mutations of isocitrate dehydrogenase (IDH) 1 and 2 occur in low­grade gliomas, acute myeloid leukemias and other types of solid cancer. By catalyzing the reversible conversion between isocitrate and α­ketoglutarate (α­KG), IDH1 and 2 contribute to the central process of metabolism, including oxidative and reductive metabolism. IDH1 and 2 mutations result in the loss of normal catalytic function and acquire neomorphic activity, facilitating the conversion of α­KG into an oncometabolite, (R)­2­hydroxyglutarate, which can cause epigenetic modifications and tumorigenesis. Small­molecule inhibitors of mutant IDH1 and 2 have been developed, and ongoing clinical trials have shown promising results in hematological malignancies, but not in gliomas. These previous findings make it necessary to identify the mechanism and develop more effective therapies for IDH1­mutant gliomas. In the present study, it was demonstrated that under hypoxic conditions, patient­derived primary glioma cells and HCT116 cells, both of which carry a monoallelic IDH1 arginine 132 to histidine mutation (R132H), have a slower growth rate than the corresponding wild­type IDH1 cells. Western blot analysis showed that IDH1 R132H­mutant cancer cells exhibited upregulated IDH2 protein expression under hypoxic conditions. Furthermore, the silencing of IDH2 using small interfering RNA significantly inhibited the growth of IDH1­mutant cells under hypoxic conditions. Finally, [U­13C5]glutamine tracer analysis showed that IDH2 knockdown reduced the reductive carboxylation of α­KG into isocitrate in HCT116R132H/+ cells under hypoxic conditions. The present study showed for the first time, to the best of our knowledge, that IDH2 plays a compensatory role in maintaining reductive carboxylation­dependent lipogenesis and proliferation in IDH1 R132H tumor cells. Therefore, IDH2 could serve as a potential anti­tumor target for IDH1­mutant tumors, which may provide a new strategy for treatment.

[RAD51 promotes proliferation and migration of glioblastoma cells and decreases sensitivity of cells to temozolomide].

Objective To investigate the role of RAD51 in cell proliferation, migration and chemosensitivity to temozolomide (TMZ) using U251 glioma cell line, and to clarify the underlying molecular mechanism. Methods TCGA database was utilized to analyze the expression changes of RAD51 in gliomas. RAD51 was over-expressed or knocked down in U251 glioma cells via lentivirus infection, or its activity was inhibited by small molecule inhibitors. Cell proliferation and migration ability were examined by CCK-8 assay, colony formation assay, and scratch wound-healing assay; CCK-8 assay and flow cytometry were performed to assess the effect of RAD51 on the sensitivity of glioma cells upon the treatment of temozolomide. Western blotting was used to determine the alteration of P53. Results The expression of RAD51 significantly increased in glioma tissues. RAD51 enhanced the proliferation and migration ability of U251 glioma cells; knockdown of RAD51 enhanced the sensitivity of U251 glioma cells to temozolomide. Over-expression of RAD51 increased the expression of P53, whereas knockdown of RAD51 decreased the expression of P53. Conclusion RAD51 plays an oncogene function in glioma cells. RAD51 over-expression enhances the proliferation and migration of glioma cells. RAD51 knockdown increases the sensitivity of glioma cells to temozolomide.

NDRG2 facilitates colorectal cancer differentiation through the regulation of Skp2-p21/p27 axis.

Poorly differentiated colorectal cancers (CRCs) are more aggressive and lack targeted therapies. We and others previously reported the predominant role of tumor-suppressor NDRG2 in promoting CRC differentiation, but the underlying mechanism is largely unknown. Herein, we demonstrate that NDRG2 induction of CRC cell differentiation is dependent on the repression of E3 ligase Skp2 activity. In patients and Ndrg2 knockout mice, NDRG2 and Skp2 are negatively correlated and associated with cell differentiation stage. Further, NDRG2 suppression of Skp2 contributes to the inductions and stabilizations of p21 and p27, which are Skp2 target proteins for degradation. The reduction of either p21 or p27 levels by shRNA can decrease NDRG2-induced AKP activity and resume cell growth inhibition, thus both p21 and p27 are required for NDRG2 effect on the promotion of cell differentiation in CRCs. The mechanistic study shows that NDRG2 suppresses ß-catenin nuclear translocation and decreases the occupancy of ß-catenin/TCF complex on Skp2 promoter, potentially through dephosphorylating GSK-3ß. By subjecting a series of NDRG2 deletion mutants to Skp2 expression, the loss of NH2-terminal domain can completely abolish NDRG2-dependent differentiation induction. Supporting the biological significance of the reciprocal relationship between NDRG2 and Skp2, an NDRG2low/Skp2high gene expression signature correlates with poor CRC patient outcome and could be considered as a diagnostic marker of CRCs.

NDRG2 acts as a PERK co-factor to facilitate PERK branch and ERS-induced cell death.

NDRG2, a newly identified tumor suppressor, is also responsive to various stresses, such as hypoxia and DNA damage. Here, we reported that in human hepatoma SK-Hep-1 and HepG2 cells, NDRG2 mRNA and protein levels were upregulated by different endoplasmic reticulum stress inducers including Tg, Tm, and DTT. Further, using NDRG2-overexpressing hepatoma cell lines and Ndrg2KO mice liver tissues, we found that, among the three branches of unfolded protein response signaling, NDRG2 facilitates protein kinase RNA-like ER kinase (PERK) pathway via interaction with PERK, enhancing its downstream ATF4 and CHOP. Functionally, NDRG2 promotes ERS-induced apoptosis partially through ATF4 or CHOP. Thus, NDRG2 is a novel ERS-responsive protein and acts as PERK co-factor to facilitate PERK branch, thereby contributing to ERS-induced apoptosis.

Immunoproapoptotic molecule scFv-Fdt-tBid modified mesenchymal stem cells for prostate cancer dual-targeted therapy.

Highly efficient target therapy is urgently needed for prostate cancer with overexpression of γ-seminoprotein (γ-SM). Recent studies indicated that mesenchymal stem cells (MSCs) are attractive candidate for cell-based, targeted therapy due to their tumor tropism. Here we designed a dual-target therapeutic system in which MSCs were engineered to produce and deliver scFv-Fdt-tBid, a novel γ-SM-targeted immunoproapoptotic molecule. Such engineered MSCs (MSC.scFv-Fdt-tBid) would home to tumor sites and release the fusion protein to induce the apoptosis of prostate cancer cells. Our data demonstrated that scFv-Fdt-tBid showed a selective, potent and dose-dependent inhibition for γ-SM-positive cells (LNCaP, C4-2, 22Rv1) rather than γ-SM-negative cells and MSCs. Importantly, MSC.scFv-Fdt-tBid caused cell death through an apoptosis-dependent manner. Further, the tropism of MSC.scFv-Fdt-tBid to prostate cancer was verified both in vitro and in vivo. Finally, the in vivo experiments demonstrated that MSC.scFv-Fdt-tBid significantly inhibited γ-SM-positive tumor growth without toxic side effects. Collectively, this study represented a novel immunoproapoptotic molecule scFv-Fdt-tBid for γ-SM-positive tumors and demonstrated the therapeutic efficiency and safety of scFv-Fdt-tBid-modified MSCs against prostate cancers.

The chimeric ubiquitin ligase SH2-U-box inhibits the growth of imatinib-sensitive and resistant CML by targeting the native and T315I-mutant BCR-ABL.

Chronic myeloid leukemia (CML) is characterized by constitutively active fusion protein tyrosine kinase BCR-ABL. Although the tyrosine kinase inhibitor (TKI) against BCR-ABL, imatinib, is the first-line therapy for CML, acquired resistance almost inevitably emerges. The underlying mechanism are point mutations within the BCR-ABL gene, among which T315I is notorious because it resists to almost all currently available inhibitors. Here we took use of a previously generated chimeric ubiquitin ligase, SH2-U-box, in which SH2 from the adaptor protein Grb2 acts as a binding domain for activated BCR-ABL, while U-box from CHIP functions as an E3 ubiquitin ligase domain, so as to target the ubiquitination and degradation of both native and T315I-mutant BCR-ABL. As such, SH2-U-box significantly inhibited proliferation and induced apoptosis in CML cells harboring either the wild-type or T315I-mutant BCR-ABL (K562 or K562R), with BCR-ABL-dependent signaling pathways being repressed. Moreover, SH2-U-box worked in concert with imatinib in K562 cells. Importantly, SH2-U-box-carrying lentivirus could markedly suppress the growth of K562-xenografts in nude mice or K562R-xenografts in SCID mice, as well as that of primary CML cells. Collectively, by degrading the native and T315I-mutant BCR-ABL, the chimeric ubiquitin ligase SH2-U-box may serve as a potential therapy for both imatinib-sensitive and resistant CML.

Chimeric ubiquitin ligases inhibit non-small cell lung cancer via negative modulation of EGFR signaling.

Targeting epidermal growth factor receptor (EGFR) represents a promising therapeutic strategy for non-small cell lung cancers (NSCLC). The ubiquitin-proteasome system (UPS) is a major pathway that mediates protein degradation. To target the degradation of EGFR, we generated two artificial ubiquitin ligases, which are composed of an EGFR-binding domain, i.e., the SH2 domain from growth factor receptor binding protein 2 (Grb2), and an ubiquitin ligase catalytic domain, i.e., the RING domain from Cbl or the U-box domain from CHIP. When the chimeric ubiquitin ligases were introduced into lung cancer SPC-A1 cells, they effectively associated with EGFR, promoted its ubiquitination and degradation, and as a result, blocked the downstream PI3K-Akt signal pathway. Moreover, cell proliferation and invasion were inhibited, the sensitivity to docetaxel-induced apoptosis was enhanced and the tumorigenicity was suppressed. In conclusion, negative modulation of EGFR signaling by the chimeric ubiquitin ligases can inhibit malignancy of SPC-A1 cells and sensitize these cells to chemotherapy, thus it may be applied to targeted therapy for NSCLC.

Two mature products of MIR-491 coordinate to suppress key cancer hallmarks in glioblastoma.

MIR-491 is commonly co-deleted with its adjacent CDKN2A on chromosome 9p21.3 in glioblastoma multiforme (GBM). However, it is not known whether deletion of MIR-491 is only a passenger event or has an important role. Small-RNA sequencing of samples from GBM patients demonstrated that both mature products of MIR-491 (miR-491-5p and -3p) are downregulated in tumors compared with the normal brain. The integration of GBM data from The Cancer Genome Atlas (TCGA), miRNA target prediction and reporter assays showed that miR-491-5p directly targets EGFR, CDK6 and Bcl-xL, whereas miR-491-3p targets IGFBP2 and CDK6. Functionally, miR-491-3p inhibited glioma cell invasion; overexpression of both miR-491-5p and -3p inhibited proliferation of glioma cell lines and impaired the propagation of glioma stem cells (GSCs), thereby prolonging survival of xenograft mice. Moreover, knockdown of miR-491-5p in primary Ink4a-Arf-null mouse glial progenitor cells exacerbated cell proliferation and invasion. Therefore, MIR-491 is a tumor suppressor gene that, by utilizing both mature forms, coordinately controls the key cancer hallmarks: proliferation, invasion and stem cell propagation.

Engineered ubiquitin ligase PTB-U-box targets insulin/insulin-like growth factor receptor for degradation and coordinately inhibits cancer malignancy.

The type 1 insulin-like growth factor receptor (IGF-1R) is a promising target for cancer therapy with antibodies and small molecule tyrosine kinase inhibitors (TKIs) which have been actively tested clinically. Evidences have demonstrated that insulin receptor (IR), which is implicated in tumorigenesis, conveys resistance to IGF-1R targeted therapy. This provided the compelling rationale for co-targeting IGF-1R and IR. Herein we have developed an approach to simultaneously down-regulate IGF-1R and IR in protein levels. By generating and screening several engineered ubiquitin ligases, we have identified that, PTB-U-box, which is composed of an IGF-1R/IR-binding domain and a functional E3 ubiquitin ligase domain, binds activated IGF-1R/IR and targets their ubiquitination and degradation. When ectopically expressed in HepG2 and HeLa cells, PTB-U-box inhibits cell proliferation and invasion, increases chemo-sensitivity, as well as interrupts glucose metabolism. Finally, intratumoral injection of adenovirus carrying PTB-U-box dramatically retards the growth of HepG2 xenograft. Therefore, well-designed engineered ubiquitin ligase represents an effective therapeutic strategy for the treatment of the cancers with co-expressed IGF-1R/IR.

Up-regulation of pVHL along with down-regulation of HIF-1α by NDRG2 expression attenuates proliferation and invasion in renal cancer cells.

The majority of renal cell carcinomas (RCCs) are characterized by loss of function of the tumor suppressor gene von Hippel Lindau (VHL), which acts as ubiquitin ligase for hypoxia-inducible factor-1α (HIF-1α). In the absence of VHL, HIF-1α protein becomes stabilized and contributes to tumorigenesis. Recent data demonstrate the antitumor efficacy of VHL promoter in RCC cells. This study demonstrates that N-Myc downstream-regulated gene 2 (NDRG2) is a potential regulator of VHL. NDRG2 is involved in proliferation and invasion of cancer cell, furthermore it is frequently down-regulated in renal cell carcinoma. Herein we evaluated the effect of NDRG2 overexpression on proliferation and invasion in human renal cancer cells. The human renal cancer cell line 786-O and A498 were infected with Ad-NDRG2 or Ad-LacZ. Overexpression of NDRG2 not only inhibited the growth of the cells, but also suppressed the invasion. Further study showed that the tumor suppressor gene VHL were up-regulated, whereas transcription factor HIF-1a and vascular endothelial growth factor (VEGF) were down-regulated in 786-O cells infected by Ad-NDRG2. Finally, in a nude mouse model, intratumoral injections of Ad-NDRG2 every 3 days for a total of seven times significantly inhibited the growth and angiogenesis of xenografted 786-O tumors. In conclusion, these data indicate that NDRG2 may be involved in proliferation and invasion by impacting the expression of VHL and HIF-1α. NDRG2 may be an attractive therapeutic target for renal cell carcinoma.