Accelerated Cell Proliferation Research Articles

Microbial Production of Retinyl Palmitate and Its Application as a Cosmeceutical

Chemically synthesized retinyl palmitate has been widely used in the cosmetic and biotechnology industry. In this study, we aimed to demonstrate the microbial production of retinyl palmitate and the benefits of microbial retinyl palmitate in skin physiology. A heterologous retinyl palmitate biosynthesis pathway was reconstructed in metabolically engineered Escherichia coli using synthetic expression modules from Pantoea agglomerans, Salinibacter ruber, and Homo sapiens. High production of retinyl palmitate (69.96 ± 2.64 mg/L) was obtained using a fed-batch fermentation process. Moreover, application of purified microbial retinyl palmitate to human foreskin HS68 fibroblasts led to increased cellular retinoic acid-binding protein 2 (CRABP2) mRNA level [1.7-fold (p = 0.001) at 100 μg/mL], acceleration of cell proliferation, and enhancement of procollagen synthesis [111% (p < 0.05) at 100 μg/mL], strongly indicating an anti-ageing-related effect of this substance. These results would pave the way for large-scale production of retinyl palmitate in microbial systems and represent the first evidence for the application of microbial retinyl palmitate as a cosmeceutical.

LncRNA HEIH accelerates cell proliferation and inhibits cell senescence by targeting miR-3619-5p/CTTNBP2 axis in ovarian cancer.

Epithelial ovarian cancer is the most lethal malignancy in gynecology. Numerous studies have confirmed that long noncoding RNAs (lncRNAs) are abnormally expressed in ovarian cancer and are closely associated with the cell proliferation and senescence in cancers. However, the role and underlying molecular mechanism of long noncoding RNA high expression in hepatocellular carcinoma (HEIH) in ovarian cancer remain unknown. Experiments including Real-time quantitative polymerase chain reaction, RNA immunoprecipitation, luciferase reporter, Fluorescence in situ hybridization, western blot, colony formation assays, β-galactosidase senescence assay, cell apoptosis, proliferation, invasion, and migration assays were applied to investigate the role of HEIH in ovarian cancer. The data were expressed as the mean ± standard deviation. Student t test was used to compare the data between two groups. The one-way analysis of variance was applied to compare the data among multiple groups with Tukey post hoc test. All experiments were repeated three times. P < 0.05 was considered statistically significant. Herein, HEIH expression was found to be up-regulated in ovarian cancer tissues (n = 25; twofold higher than normal tissues, P < 0.05) and cell lines (sixfold higher than normal ovarian epithelial cell line on average, P < 0.05), and high HEIH expression predicted poor prognosis (survival rate is about 25% after 40 mo; P < 0.05). Moreover, we found that HEIH accelerated proliferation, migration, and invasion, whereas inhibited cell senescence in ovarian cancer (P < 0.05). In mechanism, HEIH was confirmed to serve as a sponge for miR-3619-5p, and miR-3619-5p counteracted HEIH-mediated regulation of ovarian cancer (P < 0.05). Besides, cortactin-binding protein 2 (CTTNBP2) was found to be the downstream target of miR-3619-5p. Rescue assays validated that CTTNBP2 up-regulation significantly reversed the inhibitory effects of HEIH knockdown on ovarian cancer progression (P < 0.05). Furthermore, we found that HEIH facilitated tumor growth in vivo by regulating CTTNBP2 expression (P < 0.05). In conclusion, our research revealed that HEIH accelerated cell proliferation, migration and invasion, whereas inhibited cell senescence in ovarian cancer via targeting the miR-3619-5p/CTTNBP2 axis. These findings may be valuable for finding new therapeutic targets to improve ovarian cancer treatment.

Epigenetic inactivation of IRX4 is responsible for acceleration of cell growth in human pancreatic cancer.

Epigenetic gene silencing by aberrant DNA methylation is one of the important mechanisms leading to loss of key cellular pathways in tumorigenesis. Methyl‐CpG‐targeted transcriptional activation (MeTA) reactivates hypermethylation‐mediated silenced genes in a different way from DNA‐demethylating agents. Microarray coupled with MeTA (MeTA‐array) identified seven commonly hypermethylation‐mediated silenced genes in 12 pancreatic ductal adenocarcinoma (PDAC) cell lines. Among these, we focused on IRX4 (Iroquois homeobox 4) because IRX4 is located at chromosome 5p15.33 where PDAC susceptibility loci have been identified through genome‐wide association study. IRX4 was greatly downregulated in all of the analyzed 12 PDAC cell lines by promoter hypermethylation. In addition, the IRX4 promoter region was found to be frequently and specifically hypermethylated in primary resected PDACs (18/28: 64%). Reexpression of IRX4 inhibited colony formation and proliferation in two PDAC cell lines, PK‐1 and PK‐9. In contrast, knockdown of IRX4 accelerated cell proliferation in an IRX4‐expressing normal pancreatic ductal epithelial cell line, HPDE‐1. Because IRX4 is a sequence‐specific transcription factor, downstream molecules of IRX4 were pursued by microarray analyses utilizing tetracycline‐mediated IRX4 inducible PK‐1 and PK‐9 cells; CRYAB, CD69, and IL32 were identified as IRX4 downstream candidate genes. Forced expression of these genes suppressed colony formation abilities for both PK‐1 and PK‐9. These results suggest that DNA methylation‐mediated silencing of IRX4 contributes to pancreatic tumorigenesis through aberrant transcriptional regulation of several cancer‐related genes.

DLX6-AS1 accelerates cell proliferation through regulating miR-497-5p/SNCG pathway in prostate cancer.

Prostate cancer (PCa) has become the second leading cause of cancer-related mortality in males worldwide. Although the long noncoding RNA DLX6-AS1 has been recognized to be an oncogene in multiple cancers, the biological function and regulatory mechanism of DLX6-AS1 in prostate cancer are still obscure. In the present study, we observed that DLX6-AS1 was significantly upregulated in PCa tissues and cells. Knockdown of DLX6-AS1 inhibited PCa progression by suppressing cell proliferation and accelerating cell apoptosis. Molecular mechanism exploration indicated that DLX6-AS1 acted as a sponge for miR-497-5p and synuclein gamma (SNCG) was a downstream target gene of miR-497-5p. In addition, there was a negative correlation between DLX6-AS1 and miR-497-5p in PCa tissues. Rescue assays showed that SNCG overexpression could partially recover DLX6-AS1 knockdown-mediated inhibition of progression in PCa. Furthermore, xenograft tumor model was established to determine the role of DLX6-AS1 in PCa tumor growth and the results suggested that DLX6-AS1 could facilitate tumor growth by regulating SNCG in vivo. In conclusion, our study investigated the biological function and underlying mechanism of DLX6-AS1 in PCa and validated that DLX6-AS1 functioned as an oncogene through miR-497-5p/SNCG axis.

Deciphering the dual role and prognostic potential of PINK1 across cancer types.

Metabolic rewiring and deregulation of the cell cycle are hallmarks shared by many cancers. Concerted mutations in key tumor suppressor genes, such as PTEN, and oncogenes predispose cancer cells for marked utilization of resources to fuel accelerated cell proliferation and chemotherapeutic resistance. Mounting research has demonstrated that PTEN-induced putative kinase 1 (PINK1) acts as a pivotal regulator of mitochondrial homeostasis in several cancer types, a function that also extends to the regulation of tumor cell proliferative capacity. In addition, involvement of PINK1 in modulating inflammatory responses has been highlighted by recent studies, further expounding PINK1’s multifunctional nature. This review discusses the oncogenic roles of PINK1 in multiple tumor cell types, with an emphasis on maintenance of mitochondrial homeostasis, while also evaluating literature suggesting a dual oncolytic mechanism based on PINK1’s modulation of the Warburg effect. From a clinical standpoint, its expression may also dictate the response to genotoxic stressors commonly used to treat multiple malignancies. By detailing the evidence suggesting that PINK1 possesses distinct prognostic value in the clinical setting and reviewing the duality of PINK1 function in a context-dependent manner, we present avenues for future studies of this dynamic protein.

Long Non-Coding RNA USP2-AS1 Accelerates Cell Proliferation and Migration in Ovarian Cancer by Sponging miR-520d-3p and Up-Regulating KIAA1522.

BackgroundOvarian cancer is one of the malignant tumors attacking the female reproductive system. Currently, increasing studies have clearly determined the importance of long non-coding RNAs (lncRNAs) in various human cancers including ovarian cancer. However, the role and in-depth mechanism of ubiquitin specific peptidase 2 antisense RNA 1 (USP2-AS1) in ovarian cancer have been not reported yet.PurposeWe were absorbed into exploring the character of USP2-AS1 in ovarian cancer.MethodsRT-qPCR analysis reflected gene expression. The GEPIA database provided further evidences, and bioinformatics tools analyzed the potential molecules downstream USP2-AS1 in ovarian cancer. The changes on ovarian cancer cellular functions were assessed via EdU, TUNEL, JC-1 and transwell assays. RNA pull down, RIP and luciferase reporter assays estimated molecule interactions.ResultsUSP2-AS1 was obviously up-regulated in ovarian cancer tissues and cell lines. Inhibiting USP2-AS1 had anti-proliferation, pro-apoptosis, and anti-migration effects on ovarian cancer cells. Furthermore, we confirmed that USP2-AS1 sequestered miR-520d-3p to enhance KIAA1522. In addition, miR-520d-3p silence reversed the effect of depleted USP2-AS1 on ovarian cancer cellular behaviors, while such reversion was then abolished by KIAA1522 knockdown.ConclusionUSP2-AS1 facilitated ovarian cancer progression via miR-520d-3p/KIAA1522 axis, implying USP2-AS1 as a new perspective for the treatment of ovarian cancer.

Structure of CYRI-B (FAM49B), a key regulator of cellular actin assembly.

In eukaryotes, numerous fundamental processes are controlled by the WAVE regulatory complex (WRC) that regulates cellular actin polymerization, crucial for cell motility, cell-cell adhesion and epithelial differentiation. Actin assembly is triggered by interaction of the small GTPase Rac1 with CYFIP1, a key component of the WRC. Previously known as FAM49B, CYRI-B is a protein that is highly conserved across the Eukaryota and has recently been revealed to be a key regulator of Rac1 activity. Mutation of CYRI-B or alteration of its expression therefore leads to altered actin nucleation dynamics, with impacts on lamellipodia formation, cell migration and infection by intracellular pathogens. In addition, knockdown of CYRI-B expression in cancer cell lines results in accelerated cell proliferation and invasiveness. Here, the structure of Rhincodon typus (whale shark) CYRI-B is presented, which is the first to be reported of any CYRI family member. Solved by X-ray crystallography, the structure reveals that CYRI-B comprises three distinct α-helical subdomains and is highly structurally related to a conserved domain present in CYFIP proteins. The work presented here establishes a template towards a better understanding of CYRI-B biological function.

Age-associated mitochondrial DNA mutations cause metabolic remodelling that contributes to accelerated intestinal tumorigenesis.

Oxidative phosphorylation (OXPHOS) defects caused by somatic mitochondrial DNA (mtDNA) mutations increase with age in human colorectal epithelium and are prevalent in colorectal tumours, but whether they actively contribute to tumorigenesis remains unknown. Here we demonstrate that mtDNA mutations causing OXPHOS defects are enriched during the human adenoma/carcinoma sequence, suggesting they may confer a metabolic advantage. To test this we deleted the tumour suppressor Apc in OXPHOS deficient intestinal stem cells in mice. The resulting tumours were larger than in control mice due to accelerated cell proliferation and reduced apoptosis. We show that both normal crypts and tumours undergo metabolic remodelling in response to OXPHOS deficiency by upregulating the de novo serine synthesis pathway (SSP). Moreover, normal human colonic crypts upregulate the SSP in response to OXPHOS deficiency prior to tumorigenesis. Our data show that age-associated OXPHOS deficiency causes metabolic remodelling that can functionally contribute to accelerated intestinal cancer development.

Oncogenic LncRNA CASC9 in Cancer Progression.

Long non-coding RNAs (LncRNAs), with the length of over 200 nucleotides, that originate from intergenic, antisense, or promoter-proximal regions, are a large family of RNAs that lack coding capacity. Emerging evidences illustrated that LncRNAs played significant roles in a variety of cellular functions and biological processes in profuse human diseases, especially in cancers. Cancer susceptibility candidate 9 (CASC9), as a member of the LncRNAs group, firstly found its oncogenic function in esophageal cancer. In the following recent studies, a growing amount of human malignancies are verified to be correlated with CASC9, most of which are derived from the squamous epithelium tissue. This present review attempts to highlight the latest insights into the expression, functional roles, and molecular mechanisms of CASC9 in different human malignancies. In this review, the latest findings related to the pathophysiological processes of CASC9 in human cancers were summarized and analyzed, and the associated studies collected in systematic retrieval of PubMed used lncRNA and CASA9 as keywords. CASC9 expression is identified to be aberrantly elevated in a variety of malignancies. The over-expression of CASC9 has been suggested to accelerate cell proliferation, migration, cell growth and drug resistance of cancer cells, while depressing cell apoptosis, revealing its role as an oncogene. Moreover, the current review demonstrated CASC9 as closely related to the neoplastic transformation of squamous epithelial cells and squamous metaplasia in non-squamous epithelial tissues. Finally, we discuss the limitations and tremendous diagnostic/ therapeutic potential of CASC9 in various human cancers. Long non-coding RNA CASC9 likely serve as useful disease biomarkers or therapeutic targets which be effectively applied in the treatment of different kinds of cancers.

Long non-coding RNA AGAP2-AS1 accelerates cell proliferation, migration, invasion and the EMT process in colorectal cancer via regulating the miR-4,668-3p/SRSF1 axis.

Colorectal cancer (CRC) is a frequently occurring tumor. Although a number of long noncoding RNAs have been researched in CRC, the expression, function and mechanism of AGAP2-AS1 remains poorly investigated. Gene expression was analyzed by a quantitative reverse transcriptase-polymerase chain rreaction and western blot analyses. Cell counting kit-8, colony formation and Transwell assays were utilized to explore the functional role of AGAP2-AS1 in CRC. Luciferase reporter, RNA pull down and RNA immunoprecipitation assays were implemented to verify relationships between RNA molecules. In the present study, AGAP2-AS1 was unveiled as highly expressed in CRC cell lines compared to normal cells. AGAP2-AS1 knockdown suppressed cell proliferation, migration, invasion and the epithelial-to-mesenchymal transition process. Interestingly, AGAP2-AS1 sponges miR-4,668-3p to release SRSF1 in CRC. Furthermore, in the rescue functional assay, miR-4,668-3p down-regulation exacerbated the malignant behaviors of AGAP2-AS1-depleted CRC cells, whereas such effects were further offset by SRSF1 knockdown. AGAP2-AS1 facilitates cell proliferation, motility and EMT in CRC via targeting the miR-4,668-3p/SRSF1 axis. AGAP2-AS1 or SRSF1 may have potential as underlying therapeutic targets for CRC patients.

CircRAE1 promotes colorectal cancer cell migration and invasion by modulating miR-338-3p/TYRO3 axis

BackgroundGrowing evidence has revealed the involvement of circular RNAs (circRNAs) in numerous carcinogenesis. However, the role of circRNAs in the cancer biology of colorectal cancer (CRC) remains vague.MethodsQuantitative RT-PCR was used to detect the expression level of circRAE1 in CRC tissues and CRC cell lines. Cell proliferation, migration, and invasion were detected using CCK8 assay, Colony formation assay, wound-healing and Transwell assays. The interaction between circRAE1 and miR-338-3p and TRYO3 was confirmed using dual-luciferase reporter assays.ResultsWe uncovered a novel circRNA Hsa_circ_0060967 (also known as circRAE1) that was remarkably increased in CRC tissues. The high circRAE1 level was positively associated with advanced tumor stage, lymph node metastasis, and tumor size. The loss-of-function assay showed that circRAE1 accelerated cell proliferation, migration, and invasion. Besides, miR-338-3p was lowly expressed in the CRC tissues and CRC cell lines. The dual-luciferase reporter assays showed that circRAE1 could sponge miR-338-3p, which targeted TRYO3 in CRC cells. Furthermore, the overexpression of circRAE1 could rescue the impaired migration and invasion triggered by miR-338-3p mimics or si-TYRO3 in CRC cells and vice versa.ConclusionWe identified the network of circRAE1, miR-338-3p, and TYRO3 in CRC cells and determined that the increase in circRAE1 could serve as an oncogene by sponging miR-338-3p, which resulted in an upregulated TYRO3 expression. The finding suggests that circRAE1 is a potential therapeutic target and diagnostic marker for CRC treatment.

Controlled Delivery of Growth Factor by Hierarchical Nanostructured Core-Shell Nanofibers for the Efficient Repair of Critical-Sized Rat Calvarial Defect.

Electrospun nanofibers have received much attention as bone tissue-engineered scaffolds for their capacity to mimic the structure of natural extracellular matrix (ECM). Most studies have reproduced nanofibers with smooth surface for tissue engineering. This is quite different from the triple-helical nanotopography of natural collagen nanofibrils. In this study, hierarchical nanostructures were coated on the surface of drug-loaded core-shell nanofibers to mimic natural collagen nanofibrils. The nanoshish-kebab (SK) structure was decorated regularly on the surface of the nanofibers, and the inner-loaded bone morphogenetic protein 2 (BMP2) exhibited a gentle release pattern, similar to a zero-order release pattern in kinetics. The in vitro study also showed that the SK structure could accelerate cell proliferation, attachment, and osteogenic differentiation. Four groups of scaffolds were implanted in vivo to repair critical-sized rat calvarial defects: (1) PCL/PVA (control); (2) SK-PCL/PVA; (3) PCL/PVA-BMP2; and (4) SK-PCL/PVA-BMP2. Much more bone was formed in the SK-PCL/PVA group (24.57 ± 3.81%) than in the control group (1.21 ± 0.23%). The BMP2-loaded core-shell nanofibers with nanopatterned structure (SK-PCL/PVA-BMP2) displayed the best repair efficacy (76.38 ± 4.13%), followed by the PCL/PVA-BMP2 group (39.86 ± 5.74%). It was believed that the hierarchical nanostructured core-shell nanofibers could promote osteogeneration and that the SK structure showed synergistic ability with nanofiber-loaded BMP2 in vivo for bone regeneration. Thus, this BMP2-loaded core-shell nanofiber scaffold with hierarchical nanostructure holds great potential for bone tissue engineering applications.

LncRNA LINC00467 acted as an oncogene in esophageal squamous cell carcinoma by accelerating cell proliferation and preventing cell apoptosis via the miR-485-5p/DPAGT1 axis.

Esophageal carcinoma has been regarded as one of the top 10 common malignancies globally. Esophageal squamous cell carcinoma (ESCC) is an important subtype of esophageal carcinoma with approximately 20% survival rate. Long noncoding RNAs were documented to regulate the occurrence or progression of several tumors. However, neither the biological role nor the molecular mechanism of LINC00467 has been explored. This research is aimed to investigating the regulatory mechanism of LINC00467 in ESCC. In this study, a series of experiments including reverse transcription-quantitative polymerase chain reaction, Cell Counting Kit-8, luciferase reporter, western blot, and RNA immunoprecipitation were designed and conducted to explore the potential function and mechanism of LINC00467 in ESCC. According to experimental results, we found out upregulated LINC00467 improved cell proliferation, but hindered cell apoptosis. In mechanism, miR-485-5p was predicted, screened out, and validated to combine with LINC00467, which displayed lower expression in ESCC. Additionally, miR-485-5p negatively regulated and directly targeted DPAGT1. Rescue assays suggested that DPAGT1 amplification was able to recover the influence of LINC00467 deficiency on cell proliferation and apoptosis. Furthermore, knockdown of LINC00467 suppressed tumor growth in vivo. We proved that LINC00467 acted as an oncogene in ESCC by accelerating cell proliferation and preventing cell apoptosis via miR-485-5p/DPAGT1 axis. This may provide a potential diagnostic marker for ESCC treatment.

LncRNA MCF2L-AS1 aggravates proliferation, invasion and glycolysis of colorectal cancer cells via the crosstalk with miR-874-3p/FOXM1 signaling axis

Long non-coding RNAs (lncRNAs) regulate a series of biological processes, and their anomalous expression exerts critical roles in progression of multiple malignancies, including colorectal cancer (CRC). The present study was designed to provide new ideas and perspectives for the role of lncRNA MCF2L-AS1 and disclose the underlying mechanism in CRC. Herein, we observed that MCF2L-AS1 expression was enriched in CRC tissues and cell lines. Additionally, silencing of MCF2L-AS1 dramatically impeded cell proliferation, invasion and migration capacities of CRC, and distinctly attenuated the expression of invasion associated targets MMP-2 and MMP-9. Moreover, depletion of MCF2L-AS1 apparently restricted the glucose consumption and lactate production, and downregulated GLUT1 and LDHA expression. More importantly, we predicted and verified that MCF2L-AS1 acted as a molecular sponge for miR-874-3p and inversely regulated miR-874-3p expression. Interesting, FOXM1 was identified as direct target of miR-874-3p, and positively modulated by MCF2L-AS1 through sponging miR-874-3p. Mechanistically, MCF2L-AS1 accelerated cell proliferation, invasion and glycolysis through competitively binding to miR-874-3p, leading to enhance FOXM1 expression. Collectively, these outcomes highlighted that MCF2L-AS1 acted as a motivator by modulating the miR-874-3p/FOXM1 axis, thereby aggravating tumorigenesis and glycolysis progress of CRC, disclosing that MCF2L-AS1 may serve as a valuable and promising therapeutic strategy for CRC.

Long non‑coding RNA HOXA11‑AS accelerates cell proliferation and epithelial‑mesenchymal transition in hepatocellular carcinoma by modulating the miR‑506‑3p/Slug axis.

Hepatocellular carcinoma (HCC) is an aggressively malignant type of cancer with a complex pathogenesis. Multiple studies have identified that lncRNA HOXA11-AS is involved in the development of HCC. Nevertheless, the pathological mechanisms of HOXA11-AS in the development of HCC require further investigation. In the present study, the role and underlying mechanisms of HOXA11-AS in HCC were examined. RT-qPCR revealed that HOXA11-AS expression was increased, while that of miR-506-3p was decreased in HCC tissues and cells compared with that in adjacent non-tumor tissues and normal hepatic cells. Dual-luciferase reporter assay and RNA pull-down assay indicated that HOXA11-AS directly interacted with miR-506-3p. miR-506-3p downregulation reversed the inhibitory effects of HOXA11-AS deletion on cell proliferation, invasion and epithelial-mesenchymal transition (EMT), as shown by CCK-8 and Transwell assays, as well as western blot analysis. Bioinformatics analysis and dual-luciferase reporter assay indicated that Slug was a target gene of miR-506-3p. The overexpression of Slug reversed the effects of HOXA11-AS deletion on the viability, invasion and the EMT of HCC cells. Taken together, the present study demonstrates that HOXA11-AS functions as an oncogene to promote the progression of HCC via the miR-506-3p/Slug axis, providing a therapeutic target for patients with HCC.

DSCAM-AS1 accelerates cell proliferation and migration in osteosarcoma through miR-186-5p/GPRC5A signaling.

Osteosarcoma (OS) is one of the most primary bone malignancies, often occurring in adolescents or children. Numerous scientific findings have introduced that long noncoding RNAs (lncRNAs) can be involved in tumor occurrence and development. Although DSCAM-AS1 has been studied in several cancers, its role and mechanism in OS are poorly understood. In this work, high level of DSCAM-AS1 was validated in OS cell lines. Depleting DSCAM-AS1 inhibited cell proliferation, migration and EMT process in OS. Subsequently, we disclosed that DSCAM-AS1 was mainly observed in the cytoplasm of OS cells and could bind with miR-186-5p in OS. Moreover, inhibiting miR-186-5p rescued the impact of silenced DSCAM-AS1 on OS progression. Additionally, GPRC5A was verified as the target downstream of miR-186-5p, and it was negatively modulated by miR-186-5p but positively regulated by DSCAM-AS1. More importantly, DSCAM-AS1 enhanced GPRC5A level in OS by sequestering miR-186-5p. Finally, up-regulating GPRC5A reversed the influences of DSCAM-AS1 repression on the oncogenic behaviors of OS cells. Knockdown of DSCAM-AS1 suppressed NPC tumor growth in vivo. All findings uncovered that DSCAM-AS1 aggravated OS progression through sponging miR-186-5p to up-regulate GPRC5A expression. Thus, we proposed DSCAM-AS1 as a probable target for OS treatment.

Abstract 2612: Neuropilin-2 promotes enzalutamide-resistant neuroendocrine prostate cancer

Abstract Background: Neuroendocrine prostate cancer (NEPC) is a lethal subtype of prostate cancer (PC) that emerges after androgen deprivation therapy in advanced PC patients, with incidence significantly increased in recent years due to the widespread use of highly potent antiandrogens such as enzalutamide (ENZ). This study investigated the role of neuropilin-2 (NRP2) in the development and progression of ENZ-induced NEPC. Methods: Human PC tissue microarrays with different characteristics, including hormone naïve, castration resistant (CR) and neuroendocrine (NE), were analyzed for NRP2 expression by immunohistochemistry. Human PC cells were manipulated for stable knockdown or overexpression of NRP2, and then subjected to cell proliferation, cell colony formation and tumorsphere formation assays, and Western blot/RT-qPCR examination of NE marker expression. Results: NRP2 expression was elevated in CRPC and NEPC patient samples compared to hormone-naïve counterparts, which was supported by association of higher NRP2 expression with hormone-refractory and NE status of PC from additional clinical data sets. Increased NRP2 expression was found in LNCaP cells treated with ENZ (10 µM, up to 7 days) as well as in the ENZ-resistant C4-2BENZR cells (with acquired resistance to ENZ at 20 µM). Both cell lines exhibited treatment-induced NEPC features, including NE differentiation cell morphology, AR suppression and NE marker induction. Silencing of NRP2 in C4-2BENZR and 22Rv1 (with inherent ENZ resistance and NE traits) cells decreased cell proliferation by 50% and 75% respectively, which was corroborated in colony formation and tumorsphere assays. Silencing of NRP2 also increased apoptosis in both cell lines. NRP2 silencing further led to reduced expression of NE markers, including CHGA, SYP, CD56 and SOX2. Conversely, enforced expression of NRP2 in LNCaP and 22Rv1 cells accelerated cell proliferation by up to 4-fold, which was paralleled by increased cell colony and tumorsphere formation, reduced cell apoptosis, upregulation of NE marker expression (CHGA, SYP, NSE, CD56 and SOX2), and downregulation of AR and AR target genes (PSA, TMPRSS2, FKBP5 and KLK2). Mechanistically, NRP2 induced phosphorylation of STAT3 in C4-2BENZR and 22Rv1 cells as revealed by phospho-kinase antibody arrays, which will be further investigated for downstream effectors mediating NRP2's function in NEPC. Conclusion: NRP2 contributes to PC disease progression towards a NE phenotype following ENZ treatment and subsequent emergence of ENZ resistance, which provides new insights into understanding the mechanism underlying development of antiandrogen-induced NEPC and developing a new target for treating this lethal disease. Citation Format: Jing Wang, Jingjing Li, Boyang Wu, Lijuan Yin. Neuropilin-2 promotes enzalutamide-resistant neuroendocrine prostate cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2612.

MiR-942-5p promotes the proliferation and invasion of human melanoma cells by targeting DKK3

Objective The purpose of this study was to figure out the dysregulation of miR-942-5p in melanoma and its role in melanoma pathogenesis. Methods Quantitative real-time PCR (qRT-PCR) assay was used to determine the change of RNA expression. Protein expression was examined by Western blotting. miRNA target was validated through TargetScan and luciferase assay. Cell migration and invasion were detected by wound healing and transwell assay, respectively. Results Results of qRT-PCR manifested miR-942-5p were upregulated in melanoma cell. High expression of miR-942-5p in melanoma patients presented a poor prognosis. Upregulation of miR-942-5p accelerated cell proliferation, migration, and invasion in melanoma cells. Cell apoptosis was inhibited by miR-942-5p mimics. Suppression of miR-942-5p by its inhibitor showed the opposite effects in melanoma cells. TargetScan and luciferase assay showed that miR-942-5p directly targeted to the 3′-untranslated region (3′-UTR) of DKK3. Overexpression of DKK3 inhibited GSK-3β phosphorylation and reduced the expression of β-catenin in both cytoplasm and nucleus, which were induced by miR-942-5p mimics leading to the activation of Wnt/β-catenin pathway. Conclusion Upregulation of miR-942-5p was observed in melanoma cells and tissues and significantly associated with a poor prognosis. Though targeting 3′-UTR of DKK3, miR-942-5p could activate Wnt/β-catenin pathway, resulting in melanoma cell proliferation, migration, and invasion, which promoted the development of melanoma. These results showed that miR-942-5p might be a diagnosis and prognosis biomarker in melanoma.

Long noncoding RNA SNHG14 regulates ox-LDL-induced atherosclerosis cell proliferation and apoptosis by targeting miR-186-5p/WIPF2 axis.

To investigate the role of small nucleolus RNA host gene 14 (SNHG14) in the progression of atherosclerosis (AS), bioinformatics analysis, and other relevant experiments (cell counting kit-8, flow cytometry, quantitative real-time polymerase chain reaction, luciferase reporter, RNA immunoprecipitation, RNA pull-down, and western blot assays) were done. The current study revealed that SNHG14 level was high in the serum of AS patients and oxidized low-density lipoprotein (ox-LDL)-induced AS cell lines. Besides, we found that SNHG14 accelerated cell proliferation while inhibited cell apoptosis in ox-LDL-induced AS cell lines. Next, SNHG14 was confirmed to be a sponge for miR-186-5p in AS cells, and it was validated that SNHG14 regulated AS cell proliferation and apoptosis by sponging miR-186-5p. Moreover, we uncovered that WAS-interacting protein family member 2 (WIPF2) was a downstream target of miR-186-5p in AS cells. Finally, it was demonstrated that miR-186-5p modulated AS cell proliferation and apoptosis via targeting WIPF2. To conclude, our research disclosed that SNHG14 affected ox-LDL-induced AS cell proliferation and apoptosis through miR-186-5p/WIPF2 axis, which may provide a theoretical basis for the treatment and diagnosis of AS.

Transcriptomic Profile of Lymphovascular Invasion, a Known Risk Factor of Pancreatic Ductal Adenocarcinoma Metastasis.

Lymphovascular invasion (LVI) is an aggressive pathologic feature and considered a risk factor for distant metastasis. We hypothesized that pancreatic ductal adenocarcinomas (PDACs) with LVI are associated with shorter survival, as well as aggressive cancer biology and lymphangiogenesis in transcriptomic analysis. Utilizing the cancer genome atlas (TCGA)-PDAC cohort, we found that positive LVI was significantly associated with positive perineural invasion (PNI) (p = 0.023), and higher American Joint Committee on Cancer (AJCC) T (p = 0.017) and N (p < 0.001) categories. Furthermore, positive LVI was associated with shorter overall survival (OS) (p = 0.014) and was an independent risk factor of poor OS. Although there was no association between LVI status and lymphangiogenesis, epithelial-mesenchymal transition (EMT), or metastasis-related genes, Gene Set Enrichment Analysis revealed a strong association with cell-proliferation-related gene sets such as mitotic spindles (Normalized enrichment score (NES) = 1.76, p = 0.016) and G2/M checkpoints (NES = 1.75, p = 0.036), as well as with transforming growth factor beta (TGF-beta) signaling (NES = 1.61, p = 0.043), which is a known mechanism of metastasis in PDACs. In conclusion, positive LVI was an independent risk factor of poor OS in PDACs. We found that PDACs with LVI were possibly associated with accelerated cell proliferation and enhanced TGF-beta signaling independent of lymphangiogenesis. Transcriptomic profiling elucidates more precise tumor biology of LVI-positive PDACs.