Perioperative complication incidence and risk factors for retroperitoneal neuroblastoma in children: analysis of 571 patients.

BACKGROUND: Surgery plays an important role in the treatment of neuroblastoma. Perioperative complications may impact the course of neuroblastoma treatment. To date, comprehensive analyses of complications and risk factors have been lacking. METHODS: Patients with retroperitoneal neuroblastoma undergoing tumor resection were retrospectively analyzed between 2014 and 2021. The data collected included clinical characteristics, operative details, operative complications and postoperative outcomes. Risk factors for perioperative complications of retroperitoneal neuroblastoma were analyzed. RESULTS: A total of 571 patients were enrolled in this study. Perioperative complications were observed in 255 (44.7%) patients. Lymphatic leakage (28.4%), diarrhea (13.5%), and injury (vascular, nerve and organ; 7.5%) were the most frequent complications. There were three operation-related deaths (0.53%): massive hemorrhage (n = 1), biliary tract perforation (n = 1) and intestinal necrosis (n = 1). The presence of image-defined risk factors (IDRFs) [odds ratio (OR) = 2.09, P < 0.01], high stage of the International Neuroblastoma Risk Group staging system (INRGSS) (OR = 0.454, P = 0.04), retroperitoneal lymph node metastasis (OR = 2.433, P = 0.026), superior mesenteric artery encasement (OR = 3.346, P = 0.003), and inferior mesenteric artery encasement (OR = 2.218, P = 0.019) were identified as independent risk factors for perioperative complications. CONCLUSIONS: Despite the high incidence of perioperative complications, the associated mortality rate was quite low. Perioperative complications of retroperitoneal neuroblastoma were associated with IDRFs, INRGSS, retroperitoneal lymph node metastasis and vascular encasement. Patients with high-risk factors should receive more serious attention during surgery but should not discourage the determination to pursue total resection of neuroblastoma. Video Abstract (MP4 94289 KB).

Dihydromyricetin attenuates cisplatin-induced acute kidney injury by reducing oxidative stress, inflammation and ferroptosis.

BACKGROUND: Cisplatin is effective against various types of cancers. However, its clinical application is limited owing to its adverse effects, especially acute kidney injury (AKI). Dihydromyricetin (DHM), a flavonoid derived from Ampelopsis grossedentata, has varied pharmacological activities. This research aimed to determine the molecular mechanism for cisplatin-induced AKI. METHODS: A murine model of cisplatin-induced AKI (22 mg/kg, I.P.) and a HK-2 cell model of cisplatin-induced damage (30 µM) were established to evaluate the protective function of DHM. Renal dysfunction markers, renal morphology and potential signaling pathways were investigated. RESULTS: DHM decreased the levels of renal function biomarkers (blood urea nitrogen and serum creatinine), mitigated renal morphological damage, and downregulated the protein levels of kidney injury molecule-1 and neutrophil gelatinase-associated lipocalin. It upregulated the expression levels of antioxidant enzymes (superoxide dismutase and catalase expression), nuclear factor-erythroid-2-related factor 2 (Nrf2) and its downstream proteins, including heme oxygenase-1 (HO-1), glutamate-cysteine ligase catalytic (GCLC) and modulatory (GCLM) subunits, thus eventually reducing cisplatin-induced reactive oxygen species (ROS) production. Moreover, DHM partially inhibited the phosphorylation of the active fragments of caspase-8 and -3 and mitogen-activated protein kinase and restored glutathione peroxidase 4 expression, which attenuated renal apoptosis and ferroptosis in cisplatin-treated animals. DHM also mitigated the activation of NLRP3 inflammasome and nuclear factor (NF)-κB, attenuating the inflammatory response. In addition, it reduced cisplatin-induced HK-2 cell apoptosis and ROS production, both of which were blocked by the Nrf2 inhibitor ML385. CONCLUSIONS: DHM suppressed cisplatin-induced oxidative stress, inflammation and ferroptosis probably through regulating of Nrf2/HO-1, MAPK and NF-κB signaling pathways.

N6-methyladenosine modified LINC00901 promotes pancreatic cancer progression through IGF2BP2/MYC axis.

Accumulating evidence indicates that RNA methylation at N6-methyladenosine (m6A) plays an important regulatory role in gene expression and aberrant mRNA m6A modification is often associated with a variety of cancers. However, little is known whether and how m6A-modification impacts long non-coding RNA (lncRNA) and lncRNA-mediated tumorigenesis, particularly in pancreatic ductal adenocarcinoma (PDAC). In the present study, we report that a previously uncharacterized lncRNA, LINC00901, promotes pancreatic cancer cell growth and invasion and moreover, LINC00901 is subject to m6A modification which regulates its expression. In this regard, YTHDF1 serves as a reader for the m6A modified LINC00901 and downregulates the LINC00901 level. Notably, two conserved m6A sites in LINC00901 are critical to the recognition of LINC00901 by YTHDF1. Finally, RNA sequencing (RNA-seq) and gene function analysis revealed that LINC00901 positively regulates MYC through upregulation of IGF2BP2, a known RNA binding protein that can enhance MYC mRNA stability. Together, our results suggest that there is a LINC00901-IGF2BP2-MYC axis through which LINC00901 promotes PDAC progression in an m6A dependent manner.

Roles of lncRNAs in childhood cancer: Current landscape and future perspectives.

According to World Health Organization (WHO), cancer is the leading cause of death for children and adolescents. Leukemias, brain cancers, lymphomas and solid tumors, such as neuroblastoma, ostesarcoma and Wilms tumors are the most common types of childhood cancers. Approximately 400,000 children and adolescents between the ages of 0 and 19 are diagnosed with cancer each year worldwide. The cancer incidence rates have been rising for the past few decades. Generally, the prognosis of childhood cancers is favorable, but the survival rate for many unresectable or recurring cancers is substantially worse. Although random genetic mutations, persistent infections, and environmental factors may serve as contributing factors for many pediatric malignancies, the underlying mechanisms are yet unknown. Long non-coding RNAs (lncRNAs) are a group of transcripts with longer than 200 nucleotides that lack the coding capacity. However, increasing evidence indicates that lncRNAs play vital regulatory roles in cancer initiation and development in both adults and children. In particular, many lncRNAs are stable in cancer patients' body fluids such as blood and urine, suggesting that they could be used as novel biomarkers. In support of this notion, lncRNAs have been identified in liquid biopsy samples from pediatric cancer patients. In this review, we look at the regulatory functions and underlying processes of lncRNAs in the initiation and progression of children cancer and discuss the potential of lncRNAs as biomarkers for early detection. We hope that this article will help researchers explore lncRNA functions and clinical applications in pediatric cancers.

Stabilization of UCA1 by N6-methyladenosine RNA methylation modification promotes colorectal cancer progression.

BACKGROUND: UCA1 is frequently upregulated in a variety of cancers, including CRC, and it can play an oncogenic role by various mechanisms. However, how UCA1 is regulated in cancer is largely unknown. In this study, we aimed to determine whether RNA methylation at N6-methyladenosine (m6A) can impact UCA1 expression in colorectal cancer (CRC). METHODS: qRT-PCR was performed to detect the level of UCA1 and IGF2BP2 in CRC samples. CRISPR/Cas9 was employed to knockout (KO) UCA1, METTL3 and WTAP in DLD-1 and HCT-116 cells, while rescue experiments were carried out to re-express METTL3 and WTAP in KO cells. Immunoprecipitation using m6A antibody was performed to determine the m6A modification of UCA1. In vivo pulldown assays using S1m tagging combined with site-direct mutagenesis was carried out to confirm the recognition of m6A-modified UCA1 by IGF2BP2. Cell viability was measured by MTT and colony formation assays. The expression of UCA1 and IGF2BP2 in TCGA CRC database was obtained from GEPIA ( http://gepia.cancer-pku.cn ). RESULTS: Our results revealed that IGF2BP2 serves as a reader for m6A modified UCA1 and that adenosine at 1038 of UCA1 is critical to the recognition by IGF2BP2. Importantly, we showed that m6A writers, METTL3 and WTAP positively regulate UCA1 expression. Mechanically, IGF2BP2 increases the stability of m6A-modified UCA1. Clinically, IGF2BP2 is upregulated in CRC tissues compared with normal tissues. CONCLUSION: These results suggest that m6A modification is an important factor contributing to upregulation of UCA1 in CRC tissues.

Lnc-DC promotes estrogen independent growth and tamoxifen resistance in breast cancer.

Selective estrogen receptor modulators (SERMs) such as tamoxifen have proven to be effective in the treatment of estrogen receptor (ER) positive breast cancer. However, a major obstacle for such endocrine therapy is estrogen independent growth, leading to resistance, and the underlying mechanism is not fully understood. The purpose of this study was to determine whether long non-coding RNAs (lncRNAs) are involved in regulation of estrogen independent growth and tamoxifen resistance in ER positive breast cancer. Using a CRISPR/Cas9-based SAM (synergistic activation mediator) library against a focus group of lncRNAs, we identify Lnc-DC as a candidate lncRNA. Further analysis suggests that Lnc-DC is able to reduce tamoxifen-induced apoptosis by upregulation of anti-apoptotic genes such as Bcl2 and Bcl-xL. Furthermore, Lnc-DC activates STAT3 by phosphorylation (pSTAT3Y705), and the activated STAT3 subsequently induces expression of cytokines which in turn activate STAT3, forming an autocrine loop. Clinically, upregulation of Lnc-DC is associated with poor prognosis. In particular, analysis of a tamoxifen-treated patient cohort indicates that Lnc-DC expression can predict the response to tamoxifen. Together, this study demonstrates a previously uncharacterized function of Lnc-DC/STAT3/cytokine axis in estrogen independent growth and tamoxifen resistance, and Lnc-DC may serve as a potential predictor for tamoxifen response.

IGF2BP2 regulates DANCR by serving as an N6-methyladenosine reader.

The major function of Insulin-like growth factor 2 mRNA-binding protein 2 (IGF2BP2) is to regulate cell metabolism. However, emerging evidence indicates that IGF2BP2 plays a role in cancer, but the underlying mechanism is largely unknown. Here we showed that upregulation of IGF2BP2 is associated with poor outcomes of pancreatic cancer patients and suppression of IGF2BP2 inhibits cell proliferation. We further showed that IGF2BP2 regulates lncRNA DANCR. Ectopic expression IGF2BP2 enhances, whereas knockdown (KD) or knockout (KO) of IGF2BP2 suppresses DANCR expression. Moreover, in vivo RNA precipitation and reciprocal RNA immunoprecipitation revealed that IGF2BP2 interacts with DANCR. DANCR promotes cell proliferation and stemness-like properties. Experiments with xenograft models revealed that while ectopic expression of DANCR promotes, DANCR KO suppresses tumor growth. Mechanistically, DANCR is modified at N6-methyladenosine (m6A) and mutagenesis assay identified that adenosine at 664 of DANCR is critical to the interaction between IGF2BP2 and DANCR where IGF2BP2 serves a reader for m6A modified DANCR and stabilizes DANCR RNA. Together, these results suggest that DANCR is a novel target for IGF2BP2 through m6A modification, and IGF2BP2 and DANCR work together to promote cancer stemness-like properties and pancreatic cancer pathogenesis.

RETRACTED: lncRNA RMST Enhances DNMT3 Expression through Interaction with HuR

Large bodies of studies have shown that the CRISPR/Cas9-based library screening is a very powerful tool for the identification of gene functions. However, most of these studies have focused on protein-coding genes, and, furthermore, very few studies have used gene reporters for screening. In the present study, we generated DNA methyltransferase 3B (DNMT3B) reporter and screened a CRISPR/Cas9 synergistic activation mediator (SAM) library against a focused group of lncRNAs. With this screening approach, we identified Rhabdomyosarcoma 2-Associated Transcript (RMST) as a positive regulator for DNMT3B. This was confirmed by activation of the endogenous RMST by SAM or ectopic expression of RMST. Moreover, RMST knockout (KO) suppresses DNMT3, while rescue with RMST in the KO cells restores the DNMT3 level. Finally, RMST KO suppresses global DNA methylation, leading to the upregulation of methylation-regulated genes. Mechanistically, RMST promotes the interaction between the RNA-binding protein HuR and DNMT3B 3' UTR, increasing the DNMT3B stability. Together, these results not only provide the feasibility of a reporter system for CRISPR library screening but also demonstrate the previously uncharacterized factor RMST as an important player in the modulation of DNA methylation.

LINC00346 promotes pancreatic cancer progression through the CTCF-mediated Myc transcription.

Although multiple factors are known to contribute to pancreatic ductal adenocarcinoma (PDAC) progression, the role of long non-coding RNAs (lncRNAs) in PDAC remains largely unknown. In this study, we present data that long intergenic non-coding RNA 346 (LINC00346) functions as a promoting factor for PDAC development. We first show that LINC00346 is highly expressed in pancreatic tumor specimens as compared to normal pancreatic tissue based on interrogation of The Cancer Genome Atlas (TCGA) pancreatic adenocarcinoma dataset. Of significance, this upregulation of LINC00346 is associated with overall survival (OS) and disease-free survival (DFS), respectively. We further show that knockout (KO) of LINC00346 impairs pancreatic cancer cell proliferation, tumorigenesis, migration, and invasion ability. Importantly, these phenotypes can be restored by LINC00346 re-expression in KO cells (i.e., rescue experiment). RNA precipitation assays combined with mass spectrometry analysis indicate that LINC00346 interacts with CCCTC-binding factor (CTCF), a known transcriptional repressor of c-Myc. This interaction between LINC00346 and CTCF prevents the binding of CTCF to c-Myc promoter, relieving the CTCF-mediated repression of c-Myc. Thus, LINC00346 functions as a positive transcriptional regulator of c-Myc. Together, these results suggest that LINC00346 contributes to PDAC pathogenesis by activating c-Myc, and as such, LINC00346 may serve as a potential biomarker and therapeutic target for PDAC.

Linc-RoR promotes MAPK/ERK signaling and confers estrogen-independent growth of breast cancer.

BACKGROUND: The conversion from estrogen-dependent to estrogen-independent state of ER+ breast cancer cells is the key step to promote resistance to endocrine therapies. Although the crucial role of MAPK/ERK signaling pathway in estrogen-independent breast cancer cell growth is well established, the underlying mechanism is not fully understood. METHODS: In this study, we profiled lncRNA expression against a focused group of lncRNAs selected from lncRNA database. CRISPR/Cas9 was employed to knockout (KO) linc-RoR in MCF-7 cells, while rescue experiments were carried out to re-express linc-RoR in KO cells. Colony formation and MTT assays were used to examine the role of linc-RoR in estrogen-independent growth and tamoxifen resistance. Western blot and qRT-PCR were used to determine the change of protein and lncRNA levels, respectively. The expression of DUSP7 in clinical specimens was downloaded from Oncomine ( www.oncomine.org ) and the dataset from Kaplan-Meier Plotter ( http://kmplot.com ) was used to analyze the clinical outcomes in relation to DUSP7. RESULTS: We identified that linc-RoR functions as an onco-lncRNA to promote estrogen-independent growth of ER+ breast cancer. Under estrogen deprivation, linc-RoR causes the upregulation of phosphorylated MAPK/ERK pathway which in turn activates ER signaling. Knockout of linc-RoR abrogates estrogen deprivation-induced ERK activation as well as ER phosphorylation, whereas re-expression of linc-RoR restores all above phenotypes. Moreover, we show that the ERK-specific phosphatase Dual Specificity Phosphatase 7 (DUSP7), also known as MKP-X, is involved in linc-RoR KO-induced repression of MAPK/ERK signaling. Interestingly, linc-RoR KO increases the protein stability of DUSP7, resulting in repression of ERK phosphorylation. Clinical data analysis reveal that DUSP7 expression is lower in ER+ breast cancer samples than that in ER- breast cancer. Moreover, downregulation of DUSP7 expression is associated with poor patient survival. CONCLUSION: Taken together, these results suggest that linc-RoR promotes estrogen-independent growth and activation of MAPK/ERK pathway of breast cancer cells by regulating the ERK-specific phosphatase DUSP7. Thus, this study might help not only in establishing a role for linc-RoR in estrogen-independent and tamoxifen resistance of ER+ breast cancer, but also suggesting a link between linc-RoR and MAPK/ERK pathway.

FoxM1-mediated RFC5 expression promotes temozolomide resistance.

Although methylguanine-DNA-methyltransferase (MGMT) plays an important role in resistance to temozolomide (TMZ) in glioma, 40% of gliomas with MGMT inactivation are still resistant to TMZ. The underlying mechanism is not clear. Here, we report that forkhead box M1 (FoxM1) transcriptionally activates the expression of DNA repair gene replication factor C5 (RFC5) to promote TMZ resistance in glioma cells independent of MGMT activation. We showed that RFC5 expression is positively correlated with FoxM1 expression in human glioma cells and FoxM1 is able to transcriptionally activate RFC expression by interaction with the RFC5 promoter. Furthermore, knockdown of FoxM1 or RFC5 partially re-sensitizes glioma cells to TMZ. Consistently, thiostrepton, a FoxM1 inhibitor, in combination with TMZ significantly inhibits proliferation and promotes apoptosis in glioma cells. Taken together, these findings suggest that the FoxM1-RFC5 axis may mediate TMZ resistance and thiostrepton may serve as a potential therapeutic agent against TMZ resistance in glioma cells.

MALAT1-mediated tumorigenesis.

Functional genomics studies over the past decades have unveiled that the human genome transcribes a large amount of long noncoding RNAs (lncRNAs) which have been implicated in regulation of diverse cellular functions. Dysregulation of lncRNAs are often associated with human diseases including cancer. The metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) is among the most abundant and highly conserved lncRNAs. Emerging evidence has indicated that MALAT1 may have complex and extensive functions in the development and progression of cancer. In this review, we first update on the role of MALAT1 in tumorigenesis and then discuss possible molecular mechanisms that underline the MALAT1-mediated gene regulation, leading to cancer invasion and metastasis.

Egr-1 promotes hypoxia-induced autophagy to enhance chemo-resistance of hepatocellular carcinoma cells.

Previous studies suggest that early growth response gene-1 (Egr-1) plays an important role in hypoxia-induced drug-resistance. However, the mechanism still remains to be clarified. Herein, we investigated the role of Egr-1 in hypoxia-induced autophagy and its resulted hypoxia-driven chemo-resistance in Hepatocellular Carcinoma (HCC) cells. Our data demonstrated that Egr-1 was overexpressed in HCC tissues and cells and conferred them drug resistance under hypoxia. Mechanistically, Egr-1 transcriptionally regulated hypoxia-induced autophagy by binding to LC3 promoter in HCC cells, which resulted in resistance of HCC cells to chemotherapeutic agents; while dominant negative Egr-1 could inhibit autophagy level, and thus enhanced the sensitivity of HCC cells to chemotherapeutic agents, indicating that hypoxia-induced Egr-1 expression enhanced drug resistance of HCC cells likely through autophagy. Accordingly, it is suggested that a mechanism of hypoxia/Egr-1/autophagy axis might be involved in drug resistance in HCC.

Hypoxia stabilizes microtubule networks and decreases tumor cell chemosensitivity to anticancer drugs through Egr-1.

The hypoxic environment of solid tumor causes the tumor cells survive and which could protect them from death by facilitating resistance to therapy. Here, we provide evidence that hypoxia can increase tumor cell viability and proliferation through an Egr-1-dependant pathway. Hypoxia protected the microtubules from disassembly, and Egr-1 was colocalized with microtubules in different cell cycle stages. Knockdown of Egr-1 with its siRNA overcame the protection effect of hypoxia and increased the sensitivity of tumor cells to vinblastine under hypoxic conditions. Our results suggest a novel approach for increasing the sensitivity of tumor cells to chemotherapeutics that target microtubule assembly.

Calmodulin is essential for angiogenesis in response to hypoxic stress in endothelial cells.

Angiogenesis, the formation of new blood vessels that is regulated by hypoxia, is a critical process for the growth and spread of tumors. Multiple phases of this process, including migration, adhesion, and formation of new capillary tubes, are needed for optimal tumor growth. Here, a new regulatory function for Ca2+-CaM in the vascular endothelium is described. Ca2+-CaM activation induced by hypoxia in endothelial cells is essential for angiogenic cellular responses. Inhibition of Ca2+-CaM activity suppressed endothelial cell migration, adhesion on collagen I substrate, invasion and impaired in vitro endothelial cell differentiation into tube-like structures. We also reported that CaM is co-distributed with the actin structures in the lamellipodia in migrating cells, whereas the actin cytoskeleton rearrangement induced by hypoxia was disrupted and HIF-1 transcriptional activity was decreased when treated with CaM antagonists into cultures. These data indicate that Ca2+-CaM activation is more closely associated with the regulation of angiogenic key events, especially in response to hypoxic stress.