Down‐regulation of MYH10 driven by chromosome 17p13.1 deletion promotes hepatocellular carcinoma metastasis through activation of the EGFR pathway

Abstract Somatic copy number alterations (CNAs) are a genomic hallmark of cancers. Among them, the chromosome 17p13.1 deletions are recurrent in hepatocellular carcinoma (HCC). Here, utilizing an integrative omics analysis, we screened out a novel tumour suppressor gene within 17p13.1, myosin heavy chain 10 (MYH10). We observed frequent deletions (~38%) and significant down‐regulation of MYH10 in primary HCC tissues. Deletion or decreased expression of MYH10 was a potential indicator of poor outcomes in HCC patients. Knockdown of MYH10 significantly promotes HCC cell migration and invasion in vitro, and overexpression of MYH10 exhibits opposite effects. Further, inhibition of MYH10 markedly potentiates HCC metastasis in vivo. We preliminarily elucidated the mechanism by which loss of MYH10 promotes HCC metastasis by facilitating EGFR pathway activation. In conclusion, our study suggests that MYH10, a candidate target gene for 17p13 deletion, acts as a tumour suppressor and may serve as a potential prognostic indicator for HCC patients.


| Genomic CNA, gene expression and clinical relevance analyses based on publicly available data from HCC samples
The genomic copy number data and mRNA expression data of the Cancer Genome Atlas (TCGA)-liver hepatocellular carcinoma (LIHC) cohort (https://cance rgeno me.nih.gov/) were applied for genomic characterization of 17p13.1 and screening for novel tumour suppressor gene(s). The relative copy number (log 2 transformed) >0.3 was defined as genomic amplification, while the relative copy number (log 2 transformed) <−0.3 was considered as deletion. In addition to the TCGA dataset, the other 11 gene expression profile datasets were obtained from the HCCDB database (http://lifeo me.net/datab ase/hccdb), including the International Cancer Genome Consortium Liver Cancer-RIKEN Japan (ICGC-LIRI-JP) cohort, GSE22058, GSE36376, GSE63898, GSE76427, GSE10143, GSE25097, GSE14520, GSE46444, GSE54236 and GSE64041. The difference in MYH10 expression levels between HCCs and ANTLs was assessed by the Wilcoxon rank-sum test. p < 0.05 and log 2 (fold-change) <−0.2 was considered to be statistically significant. Survival information in TCGA-LIHC was used to analyse the clinical relevance of MYH10 loss or down-regulation. The PRISMA flow was shown in Figure S1.  (Table S1) as previously described. 18 The HCC tissues were used to genotype the genomic copy number of MYH10 by CNVplex assays, and all pairs of HCC tissues and ANTLs were applied for examining the protein expression levels of MYH10 by immunohistochemistry (IHC) assays. These newly diagnosed and untreated (chemotherapy or radiotherapy) HCC samples were pathologically confirmed and tumour free. Consents for sample collection were obtained from the HCC patients or their guardians. This study was performed under the supervision of Medical Ethical Committee of Beijing Institute of Radiation Medicine (Beijing, China).

| DNA extraction and CNA analyses by quantitative PCR (qPCR) assays
First, we extracted total DNA from the liver cell lines using Trizol Reagent (15596026, Invitrogen, USA). To determine the DNA copy numbers of MYH10, a pair of primers were designed on the basis of the intron sequences of MYH10. The average genomic content of three genes, including LTBP1 (at 2p22.2 locus), SATB1 (3p24.3) and ANO3 (11p14.3), which were confirmed to having no copy number alterations in our HCC cohorts (data not shown), was used as the internal reference. All primers were listed in Table S6. The qPCR assays were performed using SYBR FAST qPCR kit (KK4607, Kapa Biosystems, USA) on IQ5 real-time PCR system (BioRad, USA).

| Validation of CNAs by CNVplex assays
The genomic copy number of MYH10 was profiled by CNVplex ® assays (GENESKY, China). The CNVplex assays were performed as previously described. 18 Briefly, each sample was subjected to capillary electrophoresis after the ligation reaction and PCR amplification. The relative copy number of MYH10 was normalized to the average copy number of four reference gene loci, including POLR2A, POP1, RPP14 and TBX15. All probes were listed in Table S6. Clean data were obtained after GeneMapper 4.1 (ABI, USA) analysis.
The RT-qPCR assays were performed using SYBR FAST qPCR kit (KK4607, Kapa Biosystems, USA) on IQ5 real-time PCR system (BioRad, USA). The relative mRNA expression levels of SPRY2, DUSP6, DUSP4, SPRED1 and MYH10 were calculated with the 2 −ΔΔCt method and normalized to GAPDH, respectively. The primer sequences for RT-qPCR are shown in the Table S6.

| Cell lines and plasmids
The human normal liver cell line L-02 and HCC cell line HepG2

| Cell proliferation and colony formation assays
The proliferation rate of HCC cells was detected using the Cell Counting Kit-8 (CCK-8) (CK04, Dojindo, Japan) according to the manufacturer's instructions.
For colony formation assays, 1000 cells were seeded in 6-well plates per well. Two-three weeks later, the colonies were stained with 0.5% crystal violet for 25 min at room temperature and then scanned and counted. An average number of colonies were obtained from three replicates. Three replicates were set up for each experiment and three independent experiments were conducted. After incubation at 37°C for 24-48 h, cells that migrated to the other side of the insert were stained with 0.5% crystal violet. The inserts were subsequently rinsed clean and counted by the IX71 inverted microscope (Olympus, Japan). An average number of cells were obtained from three random fields of view of the chamber. All experiments were repeated three times.

| Nude mice assays
The experiments were carried out in compliance with the requirements of the Institutional Animal Care and Use Committee of the National Center for Protein Sciences at Beijing (Beijing, China).
Five-to six-week-old male nude BALB/c mice were purchased from Vital River Laboratories (Beijing, China). HepG2 cells transfected with scramble control or shRNA (shMYH10-1 and shMYH10-2 were pooled as 1:1) were diluted to 2 × 10 7 per mL. For subcutaneous tumour formation assay, 2 × 10 6 cells HepG2 cells were injected subcutaneously into each side of the mice back (n = 6). The length (L) and width (W) of the lump was measured every 4 days with callipers.
The equation V = (1/2) × L × W 2 was used to calculate the tumour volume. After 68 days, the mice were sacrificed, the tumours were stripped, and tumour weights were measured.
These cells (2 × 10 6 ) were injected into the lateral tail vein of BALB/c nude mice (n = 6). The first test was performed after 24 h with the IVIS@ Lumina II system (Caliper Life Sciences, USA). The luciferase substrate (Gold Biotech, USA) was injected intraperitoneally into the mice 10 min prior to the assay. Thereafter, the test was performed every 7 days. After 4 weeks, all mice were sacrificed, and their lung tissues were applied for haematoxylin and eosin (H&E) staining. The numbers of metastatic tumours were evaluated based on H&E staining.

| mRNA expression profiling upon MYH10 knockdown
Total RNAs were extracted from HepG2 cells stably transfected with scramble control (shCtrl) or shMYH10 (shMYH10-1 and shMYH10-2 were pooled as 1:1) using RNApure tissue&cell kit (CW0584S, CWBIO, China). Three replicates were available for both the shCtrl and shMYH10 groups. The Affymetrix GeneChip Human Gene U133 Array was used for gene expression profiling. The profiling was performed by CapitalBio Corporation (Beijing, China) following the protocol of Affymetrix (USA). Row data were processed by using Robust Multi-array Average (RMA). 19 The difference of gene expression levels between shMYH10 and shCtrl was assessed by DESeq2. 20 Adjusted p (false discovery rate [FDR]) <0.05 was considered to be statistically significant.

| Gene set enrichment analyses
Gene set enrichment analysis (GSEA) 21 was performed based on the paired groups (shMYH10 vs. shCtrl) with the genes ranked according to their log 2 fold-changes (shMYH10 vs. shCtrl). The significant gene sets were identified by the weighted Kolmogorov-Smirnov test from the MsigDB (v6). 21 False discovery rate (FDR) was calculated by the Benjamini-Hochberg method. The FDR <0.05 was considered to be statistically significant. Cytoscape and EnrichmentMap 22 were applied for the visualization and cluster of GSEA results, respectively.

| Immunohistochemistry (IHC) assays
The protein expression levels of MYH10 were examined by IHC assays in tumour tissues and adjacent non-tumour tissues from HCC patients.
The following operations were performed on the slides, including deparaffinization, dehydration and washing. After incubation with 3% H 2 O 2 for 10 min, the slides were placed in a citrate buffer (pH = 6.0) at high pressure for 2 min. The slides were then incubated overnight at 4°C in primary antibody dilution (anti-MYH10; 1:200; ab230823, Abcam, USA). After washing, the slides were treated by the GTVision™ TV Detection System (GK500710, DAKO, Denmark). Then, all the slides were stained with 3, 3-diaminobenzidine tetra-hydrochloride (DAB) and haematoxylin. Image information of the slides was captured through the Olympus BX51 microscopic/Digital Camera System (Olympus, Japan).
The overall score was obtained by scoring the proportion of positive staining tumour cells (0, none; 1, <1/100; 2, 1/100 to <1/10; 3, 1/10 to <1/3; 4, 1/3 to <2/3; and 5, >2/3) and the intensity of staining (0, none; F I G U R E 1 Genomic loss at chromosome 17p13.1 correlates with down-regulation of MYH10. (A) The characteristics of genomic loss at chromosome 17p13.1. The distribution of depletions (in blue) and amplifications (in red) of chromosome 17p13.1 from individual HCC on the basis of the copy number alteration (CNA) dataset of the Cancer Genome Atlas (TCGA)-liver hepatocellular carcinoma (LIHC) cohort (Materials and Methods). The 17p13.1 cytoband is emphasized by a red line. (B) Left, the rank of Pearson correlations between the mRNA levels of each protein-coding gene (n = 47) within the focally deleted region at 17p13.1 and the copy numbers of 17p13.1. Nineteen (in red) of 47 genes were identified as candidates with significant correlations (R > 0.2, p < 0.05). Right, the correlations between the mRNA levels of MYH10 and the copy numbers of 17p13.1. RPKM, reads per kilobase per million mapped reads. (C) Dysregulation of the 19 cis-regulated genes by 17p13.1 deletions in multiple gene expression datasets of HCC cohorts from the HCCDB database (http://lifeo me.net/datab ase/ hccdb). The genes with p < 0.05 (Wilcoxon rank-sum test) and log 2 (fold-change) <−0.2 were considered to be significantly down-regulated in HCC tissues compared to adjacent non-tumour liver tissues (ANTLs). (D) Down-regulation of MYH10 in 11 independent cohorts of HCC patients (including TCGA-LIHC, ICGC-LIRI-JP, GSE22058, GSE63898, GSE76427, GSE10143, GSE25097, GSE14520, GSE46444, GSE54236 and GSE64041). (E) Genomic alteration frequencies of MYH10 in 33 types of cancer from the TCGA database. (F) The correlations between the mRNA levels and the copy numbers of MYH10 in 33 types of cancer from the TCGA database, significant correlation with both R > 0.2 and p < 0.05; n.s., not significant. The abbreviations of cancer names are described on the TCGA (https://cance rgeno me.nih.gov/) database

| Statistical analyses
All quantification data are indicated as mean ± standard deviation (SD) from three independent experiments. Statistical differences of multiple group comparisons were analysed using two-sided Student's t test or ANOVA. Disease-free survival (DFS) ranges from the day of resection to the day of the first HCC recurrence, death or last follow-up. Overall survival (OS) ranges from the date of the surgery to death or the last follow-up. Survival analysis was performed by the Kaplan-Meier method and log-rank test. In all statistical tests, p < 0.05 was considered to be significant unless stated otherwise.
Statistical analyses were performed using R (version 3.1.2) software.

| Integrative omics analysis prioritizes MYH10 as a candidate functional target of chromosome 17p1deletion
Previous studies have found that losses of chromosome 17p13.1 were common in HCCs. 23,24 Tens of genes were within 17p13.1, and a large number of efforts have been made to illustrate the novel tumour suppressor gene(s) in this region. 12 In this study, we Notably, it was shown that MYH10 is consistently down-regulated (p < 0.05, log 2 [fold-change] <−0.2) in HCC tissues compared to adjacent non-tumour liver tissues (ANTLs) in 11 out of the 12 datasets ( Figure 1C and D, and Figure S1 and Table S3), increasing its candidacy as the functional target within this deleted region. Further, we analysed the prevalence of MYH10 deletion in 33 types of cancer from TCGA and found that the frequency of MYH10 deletion was greater than 30% in half of the cancer types ( Figure 1E). Moreover, the expression levels of MYH10 are significantly correlated with the copy numbers in these types of cancer ( Figure 1F), suggesting MYH10 deletion as a trans-cancer genomic feature.

| Genomic deletion or down-regulation of MYH10 predicts poor outcomes of HCC patients
To validate these genomic findings, we first genotyped the copy number of MYH10 in HCC tissues and ANTLs from a validation cohort consisting of 154 HCC patients (designated as VALI cohort; Table S1) by CNVplex assays. 25 It was confirmed that ~38% of HCC patients are affected by MYH10 deletion (Figure 2A). To further examine the dysregulation of MYH10 in HCC, we also detected the protein expression levels of MYH10 in the VALI cohort by IHC assays. MYH10 protein was predominantly located at cytoplasm and significantly decreased in HCC tissues compared to ANTLs (p = 2.40 × 10 −42 ; Figure 2B), especially in those patients with vascular invasion (p = 0.031; Table S1). Again, HCC tumours with MYH10 deletions also presented lower expression levels of MYH10 compared to those without MYH10 deletions (p = 0.0001; Figure 2C).
Consistent with the findings in clinical specimens, using RT-qPCR assays, we also observed that MYH10 is globally deleted or downregulated in HCC cell lines (including HepG2, SMMC7721, Huh7, HCCLM3 and MHCC97H), especially in those ones with higher metastatic capacity (HCCLM3 and MHCC97H), compared to the human hepatocyte cell line L-02 ( Figure 2D and E). Consistently, the MYH10 CNA-mRNA cis-correlation was observed in these types of cell lines (p = 0.0011; Figure 2F).
Further, we evaluated the potential of MYH10 deficiency to predict prognosis in HCC patients. In TCGA-LIHC cohort, the results showed that the genomic depletion at MYH10 locus is significantly correlated with decreased overall survival (OS) rate (Log-rank p = 0.017, HR = 1.55; Figure 2G) and disease-free survival (DFS) rate (Log-rank F I G U R E 2 MYH10 is down-regulated in HCC tissues, suggesting poor outcomes of HCC patients. (A) MYH10 is recurrently deleted in HCC tumours from the validation cohort (VALI, n = 154) determined by CNVplex assays. Three independent probes were applied to genotype the genomic copy number of MYH10. Four genes, including POLR2A, POP1, RPP14 and TBX15, were applied as the internal references for normalization. (B) Protein levels of MYH10 were analysed by immunohistochemistry (IHC) assays in the VALI cohort. P value was calculated by Wilcoxon rank-sum test. HCC, hepatocellular carcinoma tissue; ANTL, adjacent non-tumour liver tissue. (C) The protein expression levels of MYH10 in HCC tumours with MYH10 deletion or those with no deletion. p value was obtained by Wilcoxon rank-sum test. (D) The relative copy numbers of MYH10 were determined in multiple human hepatocyte cell lines, including one immortalized hepatocyte cell line (L-02) and five HCC cell lines (HepG2, SMMC7721, Huh7, HCCLM3 and MHCC97H) by quantitative PCR (qPCR) assays. Three genes, including SATB1, ANO3 and LTBP1, were used as internal reference for normalization. (E) The mRNA levels of MYH10 were determined in multiple human hepatocyte cell lines utilizing real-time quantitative PCR (RT-qPCR) assays. (F) The mRNA expression levels of MYH10 in MYH10 copy number deletion group and non-deletion group cell lines. p Value was obtained by Student's t-test. (G) Kaplan-Meier analysis for the overall survival (OS) rate (up) and disease-free survival (DFS) rate (bottom) of HCC patients in the TCGA-LIHC cohort. Patients with relative copy number (log 2 ) of MYH10 ≥−0.3 or <−0.3 in primary tumour tissues were designated as non-deletion or deletion subgroup, respectively. p Value was obtained by log-rank test. HR, hazard ratio; CI, confidence interval. (H) Kaplan-Meier analysis for OS rate (up) and DFS rate (bottom) of HCC patients in the TCGA-LIHC cohort. The MYH10 mRNA expression levels were classified by the higher two tertiles in the low expression group versus the lowest tertile representing the high expression group. p Value was calculated by log-rank test. (I) Kaplan-Meier curves for OS rate (up) and DFS rate (bottom) of HCC patients in the VALI cohort. The patients with MYH10 IHC score >3 or ≤3 in primary tumour tissues were designated as high or low expression subgroup, respectively. p Value was obtained by log-rank test p = 0.003, HR = 1.59; Figure 2G (H) Transwell assays revealed that overexpression of MYH10 decreases migration and invasion of HCCLM3 and MHCC97H cells. All quantification data are mean ± SD from three independent experiments. * p < 0.05, ** p < 0.01 and *** p < 0.001 (Student's t test). n.s., not significant  Figure 3F and G). However, overexpression of MYH10 was able to reduce the migration and invasion abilities of these two types of cells ( Figure 3H). Taken together, these results suggested that MYH10 has no effect on HCC cells proliferation, but plays suppressive roles in cells migration and invasion in vitro.  Figure 4E). Histological analyses confirmed that the mice injected with MYH10-depleted cells had more metastatic nodules in the lungs than the mice from the control group ( Figure 4F). Together, these results suggested that loss of MYH10 facilitates HCC metastasis in vivo.

| Depletion of MYH10 enhances the EGFR pathway
Next, we sought to explore the underlying mechanism that MYH10 promotes HCC cells migration, invasion and metastasis. To achieve this, we described by mRNA expression profiles the genes with altered mRNA expression in MYH10-depleted HepG2 cells compared to the control cells (Table S4). We identified a total of 442 down-regulated genes and 228 up-regulated genes ( Figure 5A Figure 5D and Table S5). These functional enrichment results are consistent with previous findings that MYH10 affects EMT and cell differentiation. 17,26,27 Notably, among them, negative regulation of EGF response/MAPKs subnetworks was tensely connected, which exhibited significant association between them (Figure 5D and E).  32 and Sprouty related EVH1 domain containing proteins (SPREDs, such as SPRED1 and SPRED2), 33 which were confirmed by RT-qPCR assays ( Figure 5G). Further, we confirmed in the TCGA-LIHC cohort that the mRNA expressions of these genes were positively correlated with MYH10 expression ( Figure 5H). Taken together, these data suggested that inhibition of EGFR pathway activation may be responsible for the tumour-suppressive effect of MYH10.

| Inhibition of EGFR was required for MYH10's tumour-suppressive function
We then explored the modulatory effect of MYH10 on the EGFR pathway in HCC cells. Indeed, we observed that knockdown of MYH10 in HepG2 and SMMC7721 cells increases the phosphorylation levels of EGFR (p-EGFR) and its major downstream cascades ERK1/2 (p-ERK1/2) and AKT (p-AKT) ( Figure 6A). In contrast, overexpression of MYH10 in HCCLM3 and MHCC97H cells significantly reduced the levels of p-EGFR, p-ERK1/2 and p-AKT ( Figure 6B).
These results suggested that MYH10 inhibits EGFR signalling in HCC cells.
We next explored whether the anti-tumorigenic function of MYH10 is dependent on the EGFR pathway in HCC cells. Therefore, we treated the MYH10-depleted HCC cells with Gefitinib, a kind of EGFR tyrosine kinase inhibitor, to investigate the changes in cell growth and migration. Consistently, MYH10 depletion exhibited no significant effects on the growth of HepG2 and SMMC7721 cells either with or without Gefitinib treatment ( Figure 6C). However, the Gefitinib treatment significantly limited the pro-migratory effects of MYH10 depletion on HepG2 and SMMC7721 cells ( Figure 6D).
Accordingly, knockdown of MYH10 in HepG2 and SMMC7721 cells was able to increase the levels of p-EGFR, p-ERK and p-AKT ( Figure 6E); however, Gefitinib treatment significantly counteracted the activation of the EGFR pathway caused by MYH10 deletion ( Figure 6E). In conclusion, these results suggest that MYH10 is highly likely to affect HCC progression by inhibiting the EGFR pathway.

| DISCUSS ION
In the present study, we clarified a novel candidate tumour sup- Our results also demonstrated that loss of MYH10 promotes HCC metastasis by enhancing the activation of the EGFR pathway.
Given the high prevalence of CNAs and their pivotal prognostic relevance in human cancers, it is important to dissect the underlying mechanisms of CNAs in cancer progression and treatment.
Loss of heterozygosity (LOH) on chromosome 17p13.1was found to be a common phenomenon in HCC. 9,34 Wang et al. 35 revealed the highest incidence of genomic imbalance at 17p13 (65%) compared to any other chromosome locations. LOH at 17p13.1 has also been observed in several other types of cancer, likewise in lung cancer and colon cancer. 11 These studies suggested that 17p13.1 plays an important role in the development of tumours, including HCC.
It is well known that the 17p13.1 region contains several tumour suppressor genes, such as TP53 11 and PHF23. 36 However, several other genes are located within the 17p13.1 region and their expressions are significantly correlated with the copy number of 17p13.1, therefore deserving to be investigated. Here, we prioritized a novel candidate tumour suppressor gene MYH10 within this depleted region through an integrative omics analysis. To our best knowledge, this is the first study to elaborate on the clinical relevance of MYH10 in HCC. We observed that MYH10 deletion occurred in ~38% of primary HCCs that drives the down-regulation of MYH10.
Furthermore, down-regulation of MYH10 was significantly associated with poor outcomes of HCC patients. Our results also indicated that MYH10 deletion is a trans-cancer genomic feature and deserves more attention on its biological and clinical relevance in multi types of cancer.
MYH10 is one of the isoforms of non-muscle myosin II, and has been known to participate in cell adhesion and migration. 14 Although several studies provide a close linkage between MYH10 and tumorigenesis, the roles of MYH10 are controversial. For instance, MYH10 could promote metastasis through accelerating initial rates of lamellar spreading in breast cancer. 15 In contrast, in nasopharyngeal carcinoma, MYH10 was down-regulated by miR-200a and was shown to inhibit cell migration and invasion. 16 Here, we revealed the antimetastatic role of MYH10 in HCC cells through loss-of-and gainof-function experimental assays. These discordances might be due to tumour heterogeneity that MYH10 exerts its role in a contextdependent manner. Further studies are warranted to explore the functional consequence of MYH10 in different types of cancer. Our findings highlighted that MYH10 is a novel tumour suppressor of F I G U R E 6 MYH10 exerts its tumour-suppressive role through inactivation of the EGFR pathway. (A) Knockdown of MYH10 triggers the activation of the EGFR pathway. The immunobands of EGFR, p-EGFR (Tyr1068), ERK1/2, p-ERK1/2 (Thr202/Tyr204), AKT and p-AKT (Ser473) were detected by immunoblotting assays in L-02, HepG2 and SMMC7721 cells. (B) Overexpression of MYH10 inhibits EGFR pathway in HCCLM3 and MHCC97H cells. (C) Knockdown of MYH10 has no pro-proliferation effect on HepG2 (top) and SMMC7721 (bottom) cells with or without Gefitinib treatment (10 µM). (D) Gefitinib treatment (10 µM) abolishes the pro-migratory effects of MYH10 depletion on HepG2 (top) and SMMC7721 (bottom) cells. (E) Gefitinib treatment (10 µM) eliminates the activation of MYH10 depletion on EGFR pathway in HepG2 (left) and SMMC7721 (right) cells. *** p < 0.001 (Student's t test). n.s., not significant HCC, providing an important supplement to the mechanism of HCC development and metastasis.
It is widely accepted that the EGFR pathway, as well as downstream networks involving MEK-ERK and PI3K-AKT, was hyperactivated and played important roles in promoting tumour metastasis of multiple cancers, including HCC. 29,37,38 Here, the EGFR pathway was identified as the leading pathway modulated by MYH10.
Further, we revealed that several negative regulators of EGFR or MAPKs, including SPRY2, DUSPs and SPREDs, [31][32][33] are downregulated by MYH10 depletion. These negative regulators acted at different cascades of the EGFR signalling pathway. For instance, SPRY2 can antagonize EGFR activity by inhibiting downstream ERK activation. 39 DUSPs often inhibit EGFR activation by blocking the PI3K-AKT signalling. 40 SPRED proteins modulate EGFR signalling by inhibiting the RAS/ERK pathway. 41 However, the aberrant expression of these negative regulators in HCC is largely unknown. Our findings suggested that MYH10 is a novel repressor of the EGFR pathway through inducing the expression of these negative regulators, thus possessing a novel therapeutic vulnerability in HCCs with EGFR hypoactivation.

| CON CLUS IONS
In conclusion, the survey revealed that, for the first time, MYH10 functions as a promising tumour suppressor driven by copy number deletion at 17p13.1 in the development of HCC. Depletion or down-regulation of MYH10 suggests worse outcomes in HCC patients. Depletion of MYH10 triggered activation of the EGFR pathway, which in turn promoted metastasis of HCC cells. More evidence is necessary to elucidate the mechanism of MYH10 in HCC, which may be beneficial to develop a novel treatment strategy for this malignancy.

CO N FLI C T O F I NTE R E S T S
No competing financial interests exist.