CircRNA EPHB4 modulates stem properties and proliferation of gliomas via sponging miR‐637 and up‐regulating SOX10

Gliomas are the most common type of primary brain tumors. CircRNA ephrin type‐B receptor 4 (circEPHB4) is a circular RNA derived from the receptor tyrosine kinase EPHB4. However, the clinical significance and the specific roles of circEPHB4 in gliomas and glioma cancer stem cells (CSC) have not been studied. Here, we found that circEPHB4 (hsa_circ_0081519) and SOX10 were up‐regulated and microRNA (miR)‐637 was down‐regulated in glioma tissues and cell lines. Consistently, circEPHB4 was positively correlated with SOX10 but negatively correlated with miR‐637. The altered expressions of these molecules were independently associated with overall survival of patients. CircEPHB4 up‐regulated SOX10 and Nestin by directly sponging miR‐637, thereby stimulating stemness, proliferation and glycolysis of glioma cells. Functionally, silencing circEPHB4 or increasing miR‐637 levels in glioma cells was sufficient to inhibit xenograft growth in vivo. In conclusion, the circEPHB4/miR‐637/SOX10/Nestin axis plays a central role in controlling stem properties, self‐renewal and glycolysis of glioma cells and predicts the overall survival of glioma patients. Targeting this axis might provide a therapeutic strategy for malignant gliomas.

Abbreviations

ANOVA

analysis of variance

circEPHB4

circRNA ephrin type‐B receptor 4

circRNA

circular RNA

HK2

hexokinase 2

mRNA

messenger RNA

miRNA

microRNA

PDK1

pyruvate dehydrogenase kinase 1

PI

propidium iodide

PKM2

pyruvate kinase M2

qRT‐PCR

quantitative real‐time polymerase chain reaction

RIP

RNA immunoprecipitation

SD

standard deviation

shcircEPHB4

short hairpin RNA specifically targeting circEPHB4

     

TGFβ promotes YAP‐dependent AXL induction in mesenchymal‐type lung cancer cells

The acquisition of chemoresistance remains a major cause of cancer mortality due to the limited accessibility of targeted or immune therapies. However, given that severe alterations of molecular features during epithelial‐to‐mesenchymal transition (EMT) lead to acquired chemoresistance, emerging studies have focused on identifying targetable drivers associated with acquired chemoresistance. Particularly, AXL, a key receptor tyrosine kinase that confers resistance against targets and chemotherapeutics, is highly expressed in mesenchymal cancer cells. However, the underlying mechanism of AXL induction in mesenchymal cancer cells is poorly understood. Our study revealed that the YAP signature, which was highly enriched in mesenchymal‐type lung cancer, was closely correlated to AXL expression in 181 lung cancer cell lines. Moreover, using isogenic lung cancer cell pairs, we also found that doxorubicin treatment induced YAP nuclear translocation in mesenchymal‐type lung cancer cells to induce AXL expression. Additionally, the concurrent activation of TGFβ signaling coordinated YAP‐dependent AXL expression through SMAD4. These data suggest that crosstalk between YAP and the TGFβ/SMAD axis upon treatment with chemotherapeutics might be a promising target to improve chemosensitivity in mesenchymal‐type lung cancer.

Abbreviations

AUC

area under the curve

AXL

AXL receptor tyrosine kinase

BCL2

B‐cell lymphoma 2

CTD2

cancer target discovery and development

CTGF

connective tissue growth factor

DEG

differentially expressed genes

DOXO

doxorubicin

EMT

epithelial–mesenchymal transition

Eto

etoposide

FDA

Food and Drug Administration

ITGB3

integrin beta‐3

MAPK

mitogen‐activated protein kinase

MMP2

matrix metalloproteinase‐2

MMP9

matrix metalloproteinase‐9

mRNA

messenger RNA

NF‐κB

nuclear factor kappa‐light‐chain‐enhancer of activated B cells

SBE

SMAD binding element

SERPINE1

serpin family E member 1

siRNA

small interfering RNA

ssGSEA

single‐sample gene set enrichment analysis

TCGA

The Cancer Genome Atlas

TGFβ

transforming growth factor beta

YAP

Yes‐associated protein

YAP8SA

mutants of inhibitory phosphorylation site at eight serine to Alanine of YAP

ZEB1

zinc finger E‐box binding homeobox 1

ZEB2

zinc finger E‐box‐binding homeobox 2

     

Circ_0008039 supports breast cancer cell proliferation,migration, invasion,and glycolysis by regulating the miR‐140‐3p/SKA2 axis

Circular RNAs (circRNAs) have been shown to modulate gene expression and participate in the development of multiple malignancies. The purpose of this study was to investigate the role of circ_0008039 in breast cancer (BC). The expression of circ_0008039, miR‐140‐3p, and spindle and kinetochore‐associated protein 2 (SKA2) was detected by qRT‐PCR. Cell viability, colony formation, migration, and invasion were evaluated using methylthiazolyldiphenyl‐tetrazolium bromide (MTT) assay, colony formation assay, and transwell assay, respectively. Glucose consumption and lactate production were measured using commercial kits. Protein levels of hexokinase II (HK2) and SKA2 were determined by western blot. The interaction between miR‐140‐3p and circ_0008039 or SKA2 was verified by dual‐luciferase reporter assay. Finally, a mouse xenograft model was established to investigate the roles of circ_0008039 in BC in vivo. We found that circ_0008039 and SKA2 were upregulated in BC tissues and cells, while miR‐140‐3p was downregulated. Knockdown of circ_0008039 suppressed BC cell proliferation, migration, invasion, and glycolysis. Moreover, miR‐140‐3p could bind to circ_0008039 and its inhibition reversed the inhibitory effect of circ_0008039 interference on proliferation, migration, invasion, and glycolysis in BC cells. SKA2 was verified as a direct target of miR‐140‐3p and its overexpression partially inhibited the suppressive effect of miR‐140‐3p restoration in BC cells. Additionally, circ_0008039 positively regulated SKA2 expression by sponging miR‐140‐3p. Consistently, silencing circ_0008039 restrained tumor growth via increasing miR‐140‐3p and decreasing SKA2. In conclusion, circ_0008039 downregulation suppressed BC cell proliferation, migration, invasion, and glycolysis partially through regulating the miR‐140‐3p/SKA2 axis, providing an important theoretical basis for treatment of BC.

Abbreviations

ANOVA

analysis of variance

BC

breast cancer

circRNAs

circular RNAs

DMSO

dimethyl sulfoxide

ECAR

extracellular acidification rate

ECL

enhanced chemiluminescence

FBS

fetal bovine serum

HK2

hexokinase II

MEGM

mammary epithelial growth medium

miR‐140‐3p

microRNA‐140‐3p

MTT

methylthiazolyldiphenyl‐tetrazolium bromide

PBS

phosphate‐buffered saline

PRKAR1B

protein kinase A regulatory subunit R1‐beta

SD

standard ± deviation

SKA2

spindle and kinetochore‐associated protein 2

     

EBP2, a novel NPM‐ALK‐interacting protein in the nucleolus,contributes to the proliferation of ALCL cells by regulating tumor suppressor p53

The oncogenic fusion protein nucleophosmin‐anaplastic lymphoma kinase (NPM‐ALK), found in anaplastic large‐cell lymphoma (ALCL), localizes to the cytosol, nucleoplasm, and nucleolus. However, the relationship between its localization and transforming activity remains unclear. We herein demonstrated that NPM‐ALK localized to the nucleolus by binding to nucleophosmin 1 (NPM1), a nucleolar protein that exhibits shuttling activity between the nucleolus and cytoplasm, in a manner that was dependent on its kinase activity. In the nucleolus, NPM‐ALK interacted with Epstein–Barr virus nuclear antigen 1‐binding protein 2 (EBP2), which is involved in rRNA biosynthesis. Moreover, enforced expression of NPM‐ALK induced tyrosine phosphorylation of EBP2. Knockdown of EBP2 promoted the activation of the tumor suppressor p53, leading to G₀/G₁‐phase cell cycle arrest in Ba/F3 cells transformed by NPM‐ALK and ALCL patient‐derived Ki‐JK cells, but not ALCL patient‐derived SUDH‐L1 cells harboring p53 gene mutation. In Ba/F3 cells transformed by NPM‐ALK and Ki‐JK cells, p53 activation induced by knockdown of EBP2 was significantly inhibited by Akt inhibitor GDC‐0068, mTORC1 inhibitor rapamycin, and knockdown of Raptor, an essential component of mTORC1. These results suggest that the knockdown of EBP2 triggered p53 activation through the Akt‐mTORC1 pathway in NPM‐ALK‐positive cells. Collectively, the present results revealed the critical repressive mechanism of p53 activity by EBP2 and provide a novel therapeutic strategy for the treatment of ALCL.

Abbreviations

ALCL

anaplastic large‐cell lymphoma

EBP2

EBNA1‐binding protein 2

IMT

inflammatory myofibroblastic tumors

mTOR

mechanistic target of rapamycin

mTORC1

mTOR complex 1

NoLS

nucleolar localization signal

NPM1

nucleophosmin 1

NPM‐ALK

nucleophosmin‐anaplastic lymphoma kinase

NSCLC

non‐small cell lung cancer

TPM3

tropomyosin 3

     

SIK2 represses AKT/GSK3β/β‐catenin signaling and suppresses gastric cancer by inhibiting autophagic degradation of protein phosphatases

Salt‐inducible kinase 2 (SIK2) is an important regulator in various intracellular signaling pathways related to apoptosis, tumorigenesis and metastasis. However, the involvement of SIK2 in gastric tumorigenesis and the functional linkage with gastric cancer (GC) progression remain to be defined. Here, we report that SIK2 was significantly downregulated in human GC tissues, and reduced SIK2 expression was associated with poor prognosis of patients. Overexpression of SIK2 suppressed the migration and invasion of GC cells, whereas knockdown of SIK2 enhanced cell migratory and invasive capability as well as metastatic potential. These changes in the malignant phenotype resulted from the ability of SIK2 to suppress epithelial–mesenchymal transition via inhibition of AKT/GSK3β/β‐catenin signaling. The inhibitory effect of SIK2 on AKT/GSK3β/β‐catenin signaling was mediated primarily through inactivation of AKT, due to its enhanced dephosphorylation by the upregulated protein phosphatases PHLPP2 and PP2A. The upregulation of PHLPP2 and PP2A was attributable to SIK2 phosphorylation and activation of mTORC1, which inhibited autophagic degradation of these two phosphatases. These results suggest that SIK2 acts as a tumor suppressor in GC and may serve as a novel prognostic biomarker and therapeutic target for this tumor.

Abbreviations

AMPK

AMP‐activated protein kinase

Co‐IP

co‐immunoprecipitation

EMT

epithelial–mesenchymal transition

GAPDH

glyceraldehyde‐3‐phosphate dehydrogenase

GC

gastric cancer

GEO

Gene Expression Omnibus

H&E

hematoxylin and eosin

IHC

immunohistochemistry

mTOR

mechanistic target of rapamycin

NC

negative control

PHLPP

PH domain leucine‐rich repeat protein phosphatase

PP2A

protein phosphatase 2A

qRT‐PCR

quantitative real‐time polymerase chain reaction

SIK2

salt‐inducible kinase 2

TCF/LEF

T cell factor/lymphoid enhancer‐binding factor

TCGA

The Cancer Genome Atlas

     

Downregulation of miR‐326 and its host gene β‐arrestin1 induces pro‐survival activity of E2F1 and promotes medulloblastoma growth

Persistent mortality rates of medulloblastoma (MB) and severe side effects of the current therapies require the definition of the molecular mechanisms that contribute to tumor progression. Using cultured MB cancer stem cells and xenograft tumors generated in mice, we show that low expression of miR‐326 and its host gene β‐arrestin1 (ARRB1) promotes tumor growth enhancing the E2F1 pro‐survival function. Our models revealed that miR‐326 and ARRB1 are controlled by a bivalent domain, since the H3K27me3 repressive mark is found at their regulatory region together with the activation‐associated H3K4me3 mark. High levels of EZH2, a feature of MB, are responsible for the presence of H3K27me3. Ectopic expression of miR‐326 and ARRB1 provides hints into how their low levels regulate E2F1 activity. MiR‐326 targets E2F1 mRNA, thereby reducing its protein levels; ARRB1, triggering E2F1 acetylation, reverses its function into pro‐apoptotic activity. Similar to miR‐326 and ARRB1 overexpression, we also show that EZH2 inhibition restores miR‐326/ARRB1 expression, limiting E2F1 pro‐proliferative activity. Our results reveal a new regulatory molecular axis critical for MB progression.

Abbreviations

ARRB1

β‐arrestin1

BTC

bulk tumor cell

CSCs

cancer stem cells

EZH2

enhancer of zeste homolog 2

GCP

granule cell progenitors

MB

medulloblastoma

OFC

oncosphere‐forming cell

     

Comprehensive cross‐platform comparison of methods for non‐invasive EGFR mutation testing: results of the RING observational trial

Several platforms for noninvasive EGFR testing are currently used in the clinical setting with sensitivities ranging from 30% to 100%. Prospective studies evaluating agreement and sources for discordant results remain lacking. Herein, seven methodologies including two next‐generation sequencing (NGS)‐based methods, three high‐sensitivity PCR‐based platforms, and two FDA‐approved methods were compared using 72 plasma samples, from EGFR‐mutant non‐small‐cell lung cancer (NSCLC) patients progressing on a first‐line tyrosine kinase inhibitor (TKI). NGS platforms as well as high‐sensitivity PCR‐based methodologies showed excellent agreement for EGFR‐sensitizing mutations (K = 0.80–0.89) and substantial agreement for T790M testing (K = 0.77 and 0.68, respectively). Mutant allele frequencies (MAFs) obtained by different quantitative methods showed an excellent reproducibility (intraclass correlation coefficients 0.86–0.98). Among other technical factors, discordant calls mostly occurred at mutant allele frequencies (MAFs) ≤ 0.5%. Agreement significantly improved when discarding samples with MAF ≤ 0.5%. EGFR mutations were detected at significantly lower MAFs in patients with brain metastases, suggesting that these patients risk for a false‐positive result. Our results support the use of liquid biopsies for noninvasive EGFR testing and highlight the need to systematically report MAFs.

Abbreviations

BEAMing

beads, emulsion, amplification, and magnetics

cfDNA

circulating free DNA, cell‐free DNA

cobas

cobas^® EGFR Mutation Test v2 (Roche Diagnostics)

ctDNA

circulating tumor DNA

CUSUM

cumulative sum

ddPCR

droplet digital polymerase chain reaction

dPCR

digital polymerase chain reaction

EGFR

epidermal growth factor receptor

FFPE

formalin‐fixed, paraffin‐embedded

ICC

intraclass correlation coefficient

MAF

mutant allele frequency

NGS platforms

Ion S5™ XL and GeneRead™

NGS

next‐generation sequencing

NSCLC

non‐small‐cell lung cancer

PNA‐Q‐PCR

peptic nucleic acid probe‐based real‐time polymerase chain reaction

Therascreen

Therascreen EGFR Plasma RGQ PCR Kit (QIAgen)

TKI

tyrosine kinase inhibitor

     

A novel circular RNA,hsa_circ_0030998 suppresses lung cancer tumorigenesis and Taxol resistance by sponging miR‐558

Circular RNAs (circRNAs) are single‐stranded RNAs which form a covalently closed continuous loop. Although originally shown to be non‐protein‐coding, some circRNAs can give rise to micropeptides. circRNAs have also been shown to play essential regulatory roles in a variety of developmental and disease processes. In a previous study, hsa_circ_0030998 was identified as a circRNA downregulated in lung cancer, but its potential implications and mechanisms in lung cancer were not addressed. Here, we showed that overexpressing circ_0030998 decreased proliferation, migration, and invasion of lung cancer cells, while also dampening resistance to Taxol, a classical antitumor drug. Depleting circ_0030998 reversed these phenotypic effects. A high circ_0030998 expression was correlated with a high survival rate in lung cancer patients. Additionally, we found circ_0030998 could downregulate miR‐558 expression, serving as a microRNA sponge. In conclusion, our data support that hsa_circ_0030998 can slow down the progression of lung cancer by targeting miR‐558 and suppress malignant phenotypes such as proliferation, migration, and invasion progression of lung cancer cells. Therefore, we highlight that circ_0030998 could be a novel tumor suppressor of lung cancer.

Abbreviations

circRNA

circular RNA

IP

immunoprecipitation

LAMP1

lysosomal‐associated membrane protein 1

miRNA

microRNA

MMP

matrix metalloproteinase

NC

negative control

     

The FABP12/PPARγ pathway promotes metastatic transformation by inducing epithelial‐to‐mesenchymal transition and lipid‐derived energy production in prostate cancer cells

Early stage localized prostate cancer (PCa) has an excellent prognosis; however, patient survival drops dramatically when PCa metastasizes. The molecular mechanisms underlying PCa metastasis are complex and remain unclear. Here, we examine the role of a new member of the fatty acid‐binding protein (FABP) family, FABP12, in PCa progression. FABP12 is preferentially amplified and/or overexpressed in metastatic compared to primary tumors from both PCa patients and xenograft animal models. We show that FABP12 concurrently triggers metastatic phenotypes (induced epithelial‐to‐mesenchymal transition (EMT) leading to increased cell motility and invasion) and lipid bioenergetics (increased fatty acid uptake and accumulation, increased ATP production from fatty acid β‐oxidation) in PCa cells, supporting increased reliance on fatty acids for energy production. Mechanistically, we show that FABP12 is a driver of PPARγ activation which, in turn, regulates FABP12''s role in lipid metabolism and PCa progression. Our results point to a novel role for a FABP‐PPAR pathway in promoting PCa metastasis through induction of EMT and lipid bioenergetics.

Abbreviations

AR

androgen receptor

ATP

adenosine triphosphate

CN

copy number

CPT1

carnitine palmitoyltransferase I

CS

citrate synthase

EMT

epithelial–mesenchymal transition

ET

electron transfer‐state

FABP

fatty acid‐binding protein

LD

lipid droplet

OA

oleic acid

PCa

prostate cancer

PPAR

peroxisome proliferator‐activated receptor

PPRE

peroxisome proliferator‐activated receptor response element

TZD

thiazolidinediones

     

Intratumoral immunosuppression profiles in 11q‐deleted neuroblastomas provide new potential therapeutic targets

High‐risk neuroblastoma (NB) patients with 11q deletion frequently undergo late but consecutive relapse cycles with fatal outcome. To date, no actionable targets to improve current multimodal treatment have been identified. We analyzed immune microenvironment and genetic profiles of high‐risk NB correlating with 11q immune status. We show in two independent cohorts that 11q‐deleted NB exhibits various immune inhibitory mechanisms, including increased CD4+ resting T cells and M2 macrophages, higher expression of programmed death‐ligand 1, interleukin‐10, transforming growth factor‐beta‐1, and indoleamine 2,3‐dioxygenase 1 (P < 0.05), and also higher chromosomal breakages (P ≤ 0.02) and hemizygosity of immunosuppressive miRNAs than MYCN‐amplified and other 11q‐nondeleted high‐risk NB. We also analyzed benefits of maintenance treatment in 83 high‐risk stage M NB patients focusing on 11q status, either with standard anti‐GD2 immunotherapy (n = 50) or previous retinoic acid‐based therapy alone (n = 33). Immunotherapy associated with higher EFS (50 vs. 30, P = 0.028) and OS (72 vs. 52, P = 0.047) at 3 years in the overall population. Despite benefits from standard anti‐GD2 immunotherapy in high‐risk NB patients, those with 11q deletion still face poor outcome. This NB subgroup displays intratumoral immune suppression profiles, revealing a potential therapeutic strategy with combination immunotherapy to circumvent this immune checkpoint blockade.

Abbreviations

11q‐del

11q‐deleted

ADCC

antibody‐dependent cellular cytotoxicity

CDC

complement‐dependent cytotoxicity

COJEC

chemotherapeutic agents cisplatin, vincristine, carboplatin, etoposide, and cyclophosphamide

CTLA‐4

cytotoxic T lymphocyte antigen 4

EFS

event‐free survival

FISH

fluorescence in situ hybridization

HR

hazard ratio

ICI

immune checkpoint inhibitor

IDO1

indoleamine 2,3‐dioxygenase 1

IFN‐γ

interferon‐γ

IL‐10

interleukin 10

INRG

International Neuroblastoma Risk Group

miR

microRNA

MLPA

multiplex ligation‐dependent probe amplification

MMR

mismatch repair

MNA

MYCN amplification

MS

metastatic special stage

MSI

microsatellite instability

NB

neuroblastoma

NCA

numerical chromosome aberrations

NOS

nitric oxide synthase

OS

overall survival

PD‐1

programmed cell death protein 1

PD‐L1

programmed death‐ligand 1

SCA

segmental chromosome aberrations

TAM

tumor‐associated macrophages

Tfh

follicular helper T cells

TGF‐β

tumor growth factor‐β

TMB

tumor mutational burden

TME

tumor microenvironment

TNF‐α

tumor necrosis factor‐α

Treg

regulatory T cells

     

Circulating tumor cell detection and single‐cell analysis using an integrated workflow based on ChimeraX®‐i120 Platform: A prospective study

Circulating tumor cell (CTC) analysis holds great potential to be a noninvasive solution for clinical cancer management. A complete workflow that combined CTC detection and single‐cell molecular analysis is required. We developed the ChimeraX^®‐i120 platform to facilitate negative enrichment, immunofluorescent labeling, and machine learning‐based identification of CTCs. Analytical performances were evaluated, and a total of 477 participants were enrolled to validate the clinical feasibility of ChimeraX^®‐i120 CTC detection. We analyzed copy number alteration profiles of isolated single cells. The ChimeraX^®‐i120 platform had high sensitivity, accuracy, and reproducibility for CTC detection. In clinical samples, an average value of > 60% CTC‐positive rate was found for five cancer types (i.e., liver, biliary duct, breast, colorectal, and lung), while CTCs were rarely identified in blood from healthy donors. In hepatocellular carcinoma patients treated with curative resection, CTC status was significantly associated with tumor characteristics, prognosis, and treatment response (all P < 0.05). Single‐cell sequencing analysis revealed that heterogeneous genomic alteration patterns resided in different cells, patients, and cancers. Our results suggest that the use of this ChimeraX^®‐i120 platform and the integrated workflow has validity as a tool for CTC detection and downstream genomic profiling in the clinical setting.

Abbreviations

ADABOOST

AdaBoost classification trees

AFP

alpha‐fetoprotein

AUC

areas under the curve

BC

breast cancer

BCLC

barcelona clinic liver cancer

BHL

benign hepatic lesion

CCD

charge‐coupled device

CHB

chronic hepatitis B

CK

cytokeratin

CNA

copy number alteration

CNLC

Chinese staging for liver cancer

CRC

colorectal cancer

CTC

circulating tumor cell

CTM

circulating tumor microemboli

CV

coefficient of variation

DAPI

4’,6‐diamidine‐2’‐phenylindole dihydrochloride

EpCAM

epithelial cell adhesion molecule

FPR

false‐positive rate

GBM

stochastic gradient boosting

HCC

hepatocellular carcinoma

HD

healthy donor

ICC

intrahepatic cholangiocarcinoma

LC

liver cirrhosis

LCA

lung cancer

LOD

limit of detection

PBS

phosphate‐buffered saline

PCR

polymerase chain reaction

RF

random forest

ROC

receiver operating characteristic

SVM

support vector machines

TCGA

The Cancer Genome Atlas

TPR

true‐positive rate

TTR

time to recurrence

WBC

white blood cell

WGA

whole‐genome amplification

WGS

whole‐genome sequencing

XGB

extreme gradient boosting

     

Long non‐coding RNA RACGAP1P promotes breast cancer invasion and metastasis via miR‐345‐5p/RACGAP1‐mediated mitochondrial fission

Long non‐coding RNAs (lncRNAs) are emerging as key molecules in various cancers, yet their potential roles in the pathogenesis of breast cancer are not fully understood. Herein, using microarray analysis, we revealed that the lncRNA RACGAP1P, the pseudogene of Rac GTPase activating protein 1 (RACGAP1), was up‐regulated in breast cancer tissues. Its high expression was confirmed in 25 pairs of breast cancer tissues and 8 breast cell lines by qRT‐PCR. Subsequently, we found that RACGAP1P expression was positively correlated with lymph node metastasis, distant metastasis, TNM stage, and shorter survival time in 102 breast cancer patients. Then, in vitro and in vivo experiments were designed to investigate the biological function and regulatory mechanism of RACGAP1P in breast cancer cell lines. Overexpression of RACGAP1P in MDA‐MB‐231 and MCF7 breast cell lines increased their invasive ability and enhanced their mitochondrial fission. Conversely, inhibition of mitochondrial fission by Mdivi‐1 could reduce the invasive ability of RACGAP1P‐overexpressing cell lines. Furthermore, the promotion of mitochondrial fission by RACGAP1P depended on its competitive binding with miR‐345‐5p against its parental gene RACGAP1, leading to the activation of dynamin‐related protein 1 (Drp1). In conclusion, lncRNA RACGAP1P promotes breast cancer invasion and metastasis via miR‐345‐5p/RACGAP1 pathway‐mediated mitochondrial fission.

Abbreviations

CDS

coding sequence

ceRNAs

competitive endogenous RNAs

Drp1

dynamin‐related protein 1

FFPE

formalin‐fixed paraffin‐embedded

lncRNAs

long non‐coding RNAs

miRNAs

microRNAs

RACGAP1

Rac GTPase activating protein 1

TCGA

The Cancer Genome Atlas

     

5‐Hydroxymethylcytosine signature in circulating cell‐free DNA as a potential diagnostic factor for early‐stage colorectal cancer and precancerous adenoma

Approximately 85% colorectal cancers (CRCs) are thought to evolve through the adenoma‐to‐carcinoma sequence associated with specific molecular alterations, including the 5‐hydroxymethylcytosine (5hmC) signature in circulating cell‐free DNA (cfDNA). To explore colorectal disease progression and evaluate the use of cfDNA as a potential diagnostic factor for CRC screening, here, we performed genome‐wide 5hmC profiling in plasma cfDNA and tissue genomic DNA (gDNA) acquired from 101 samples (63 plasma and 38 tissues), collected from 21 early‐stage CRC patients, 21 AD patients, and 21 healthy controls (HC). The gDNA and cfDNA 5hmC signatures identified in gene bodies and promoter regions in CRC and AD groups were compared with those in HC group. All the differential 5hmC‐modified regions (DhMRs) were gathered into four clusters: Disease‐enriched, AD‐enriched, Disease‐lost, and AD‐lost, with no overlap. AD‐related clusters, AD‐enriched and AD‐lost, displayed the unique 5hmC signals in AD patients. Disease‐enriched and Disease‐lost clusters indicated the general 5hmC changes when colorectal lesions occurred. Cancer patients with a confirmable adenoma history segmentally gathered in AD‐enriched clusters. KEGG functional enrichment and GO analyses determined distinct differential 5hmC‐modified profiles in cfDNA of HC individuals, AD, and CRC patients. All patients had comprehensive 5hmC signatures where Disease‐enriched and Disease‐lost DhMR clusters demonstrated similar epigenetic modifications, while AD‐enriched and AD‐lost DhMR clusters indicated complicated subpopulations in adenoma. Analysis of CRC patients with adenoma history showed exclusive 5hmC‐gain characteristics, consistent with the ‘parallel’ evolution hypothesis in adenoma, either developed through the adenoma‐to‐carcinoma sequence or not. These findings deepen our understanding of colorectal disease and suggest that the 5hmC modifications of different pathological subtypes (cancer patients with or without adenoma history) could be used to screen early‐stage CRC and assess adenoma malignancy with large‐scale follow‐up studies in the future.

Abbreviations

5hmC

5‐hydroxymethylcytosine

AD

precancerous adenoma

cfDNA

cell‐free DNA

CRC

colorectal cancer

DhmR

differential 5hmC‐modified regions

gDNA

genomics DNA

HC

healthy control

hMRs

5hmC‐modified regions