The research, led by Yue Cheng and colleagues at Tongde Hospital in Zhejiang Province, China, began with a straightforward question: which genes distinguish aggressive prostate tumours from healthy prostate tissue? Using the TIMER2.0 pan-cancer database, the team surveyed SURF6 expression across multiple cancer types and found it significantly elevated in a range of malignancies, including bladder, breast, colon, liver, and stomach cancers. Crucially, prostate adenocarcinoma was among the tumour types showing robust SURF6 upregulation. The researchers then validated this finding using data from The Cancer Genome Atlas (TCGA), immunohistochemical staining from the Human Protein Atlas, and their own patient samples—frozen tumour tissues and matched adjacent normal tissue collected from seven patients undergoing primary surgery. Both Western blotting and quantitative reverse-transcription PCR confirmed that SURF6 was consistently more abundant in tumour tissue than in normal prostate.

SURF6 is not a random suspect. It is a highly conserved nucleolar matrix protein that binds nucleic acids, participates in the early steps of pre-ribosomal RNA processing, and contributes to ribosome biogenesis and cell-cycle control. Because rapidly proliferating tumour cells demand an enormous capacity for protein synthesis, dysregulation of nucleolar proteins such as SURF6 has long been viewed as a hallmark of cancer biology. What the new study adds is direct evidence that in prostate cancer, SURF6 is not merely a passenger but an active promoter of malignant behaviour.

The clinical data lent weight to this idea. When the team stratified prostate cancer patients by SURF6 expression, several worrying patterns emerged. Tumours classified as T3 or T4—larger, more locally advanced growths—expressed significantly more SURF6 than earlier T1 and T2 stage tumours. Patients whose cancer had spread to lymph nodes (N1 stage) showed higher SURF6 levels than those without nodal involvement, and men with Gleason scores of 8 or above, indicating poorly differentiated and aggressive disease, carried higher SURF6-associated risk scores than those with scores of 6. Most strikingly, survival analysis using the GEPIA2 platform showed that patients with high SURF6 expression had significantly worse overall survival and disease-free survival than those whose tumours expressed lower levels.

To determine whether SURF6 actually drives these behaviours or merely accompanies them, the researchers manipulated its levels in prostate cancer cell lines. They used small interfering RNA to silence SURF6 in PC3 cells, an aggressive androgen-independent line, and introduced a SURF6 overexpression plasmid into DU145 cells. The results were unambiguous. CCK-8 proliferation assays and colony formation experiments showed that depleting SURF6 markedly slowed the growth of PC3 cells, while boosting SURF6 accelerated the proliferation of DU145 cells. Wound healing and Transwell invasion assays told the same story for motility: silencing SURF6 suppressed migration and invasion, whereas overexpression enhanced them.

Perhaps the most consequential discovery concerns cancer stem cells—the rare, self-renewing subpopulation of tumour cells believed to seed recurrence and resist conventional therapies. Using sphere-formation assays, a standard test of stem-like self-renewal, the researchers found that SURF6 knockdown drastically impaired the ability of PC3 cells to form tumour spheres, while SURF6 overexpression increased sphere formation in DU145 cells. Flow cytometry revealed corresponding shifts in the proportions of cells carrying CD44 and CD133, well-established surface markers of prostate cancer stem cells. Western blotting extended the picture to intracellular stemness factors: levels of SOX4, Nanog, and OCT4 all fell when SURF6 was silenced and rose when it was overexpressed. In short, SURF6 appears to help prostate cancer cells maintain the stem-like identity that makes them so difficult to eradicate.

Having established what SURF6 does, the team turned to how its expression is controlled—and here the study enters one of the most dynamic areas of modern molecular biology: epitranscriptomics. Specifically, the researchers investigated N6-methyladenosine, or m6A, the most abundant internal chemical modification on eukaryotic messenger RNA. m6A marks are installed by a “writer” complex whose catalytic core is the methyltransferase METTL3, and they are interpreted by “reader” proteins such as YTHDF1, which typically enhances the translation or stability of methylated transcripts. Dysregulated m6A signalling has been implicated in numerous cancers, but its role in prostate cancer stemness had remained incompletely understood.

The evidence that SURF6 is an m6A target built up methodically. First, the team measured global m6A levels and found them significantly higher in PC3 and DU145 prostate cancer cells than in the normal prostate epithelial line RWPE-1. Sorting the cancer cells into CD44-positive and CD44-negative fractions by fluorescence-activated cell sorting revealed that the stem-like CD44-positive population carried even higher m6A levels. A computational survey of the SURF6 messenger RNA sequence using the SRAMP prediction tool identified several high-confidence m6A sites, and RNA immunoprecipitation with an anti-m6A antibody confirmed that SURF6 transcripts are indeed methylated—with enrichment strongest in the CD44-positive fraction. TCGA correlation analysis pointed to METTL3 as the relevant writer: knocking down METTL3 in PC3 cells reduced SURF6 at both RNA and protein levels, while overexpressing METTL3 in DU145 cells increased it.

The reader protein fell into place next. YTHDF1 expression correlated with SURF6 across prostate cancer samples, and RNA immunoprecipitation with a YTHDF1 antibody pulled down SURF6 messenger RNA from both cell lines. A reciprocal RNA pull-down experiment, using biotin-labelled SURF6 RNA as bait, retrieved YTHDF1 protein, confirming a physical interaction. Functionally, silencing YTHDF1 lowered SURF6 expression, whereas YTHDF1 overexpression raised it. The decisive experiment came from dual-luciferase reporters carrying either the wild-type SURF6 3′ untranslated region or a mutant version in which the predicted m6A motif had been disrupted. METTL3 and YTHDF1 both enhanced the activity of the wild-type reporter but had little effect on the mutant, demonstrating that this specific m6A site is required for regulation. An actinomycin D chase assay added the final mechanistic detail: in the absence of YTHDF1, SURF6 messenger RNA degraded more rapidly, while YTHDF1 overexpression stabilised the transcript.

With the regulatory circuit defined, the researchers asked how SURF6 exerts its effects downstream. The answer pointed squarely at CDK4, a cyclin-dependent kinase that drives the G1-to-S transition of the cell cycle and is already the target of approved inhibitors in other cancers. SURF6 knockdown reduced CDK4 expression, and the two genes’ expression correlated significantly in TCGA prostate cancer data. Actinomycin D decay assays showed that CDK4 messenger RNA decayed faster when SURF6 was silenced, and RNA pull-down experiments demonstrated that SURF6 physically associates with CDK4 transcripts—suggesting SURF6 may stabilise CDK4 mRNA, a striking role for a protein classically known as a nucleolar component of the ribosome-building machinery.

Rescue experiments sealed the causal chain. When the researchers forced CDK4 overexpression in SURF6-depleted PC3 cells, the accumulation of cells stuck in the G0/G1 phase was relieved, and proliferation, colony formation, migration, and invasion largely recovered. Conversely, knocking down CDK4 in SURF6-overexpressing DU145 cells blunted the pro-tumorigenic effects of SURF6. The same rescue logic applied to stemness: CDK4 overexpression restored sphere formation, the CD44-positive and CD133-positive fractions, and the expression of CD44, SOX4, Nanog, and OCT4 in SURF6-silenced cells, while CDK4 knockdown reversed these gains in SURF6-overexpressing cells.

Finally, the team translated their findings into living animals. They implanted nude mice with PC3 cells engineered to express a short hairpin RNA against SURF6, alongside control cells carrying a non-targeting construct. Tumours in the SURF6-knockdown group grew significantly more slowly, with markedly smaller volumes and lower final weights. Histological examination revealed reduced Ki-67 staining, a sign of diminished proliferation, while immunofluorescence and immunohistochemistry showed lower levels of CD44 and CDK4 in the shrunken tumours—confirming in vivo that the molecular pathway identified in culture operates in actual tumour growth.

Taken together, the study outlines a complete regulatory axis: METTL3 installs m6A marks on the SURF6 transcript, YTHDF1 reads those marks and stabilises the message, elevated SURF6 sustains CDK4 expression, and CDK4 drives both cell-cycle progression and the maintenance of cancer stem cell identity. Each link in this chain represents a potential point of therapeutic intervention. Because CDK4/6 inhibitors are already in clinical use for breast cancer, the prospect of extending such approaches to SURF6-high prostate tumours is an obvious next step, as is the development of m6A-pathway inhibitors capable of turning down SURF6 at the RNA level. The authors caution that their clinical sample size was small and that further validation in large patient cohorts will be needed, but the convergence of bioinformatics, cell biology, and animal modelling makes a compelling case that a nucleolar protein once known mainly for ribosome assembly sits at the heart of prostate cancer’s most dangerous behaviours. Targeting SURF6, the researchers suggest, could one day attack both the growth and the recurrence-seeding stem cell reservoir of the disease simultaneously.

Epithelial-mesenchymal transition is a crucial biological process during cancer progression, significantly contributing to tumor invasiveness and metastasis. In this study, the authors detail how the induction of EMT facilitates the transition of adherent epithelial cells into migratory mesenchymal cells, subsequently promoting cancer cells’ ability to invade surrounding tissues. Specifically in glioblastoma, this transition is rampant and correlates with increased malignancy.

Shikonin’s mechanisms of action begin with its effects on the p53 tumor suppressor protein, a key regulator of cell cycle and apoptosis. The authors report that Shikonin elevates p53 expression, which plays a pivotal role in preventing cancer cell proliferation and survival. Enhanced levels of p53 activate downstream targets that induce apoptosis and inhibit cell growth, making it a potent agent against tumor growth.

In addition to p53, this study highlights the significance of miR-361-5p in mediating Shikonin’s anti-tumor effects. MicroRNAs are small, non-coding RNAs that regulate gene expression at the post-transcriptional level. The upregulation of miR-361-5p in glioblastoma cells treated with Shikonin leads to the suppression of ZEB1, a transcription factor fundamentally involved in promoting EMT. By reducing ZEB1 levels, Shikonin effectively removes the impetus for EMT, thereby hindering the potent migratory and invasive capabilities of glioblastoma cells.

Interestingly, the study utilizes multiple experimental approaches to confirm Shikonin’s effectiveness. The researchers employed in vitro assays with various glioblastoma cell lines to assess cell viability, migration, and invasion. In tandem, they utilized Western blot analysis and quantitative RT-PCR to measure the expressions of p53, miR-361-5p, and ZEB1, establishing a clear biochemical pathway influenced by Shikonin.

The implications of this research are monumental, providing a scientific basis for utilizing Shikonin as a viable therapeutic strategy against glioblastoma. The findings emphasize not only the potential for Shikonin as a standalone treatment but also suggest its possible integration into combination therapies, where traditional chemotherapeutic agents could be used alongside natural compounds like Shikonin.

Furthermore, the study contextualizes the significance of deriving therapies from natural products. With increasing resistance to conventional chemotherapy agents, natural compounds like Shikonin present alternative routes for treatment development. These substances often possess multi-targeted mechanisms that can effectively tackle the heterogeneous nature of tumors, such as glioblastoma.

Building on this premise, the study invites additional investigation into Shikonin’s role with other oncogenic pathways, particularly those associated with tumor microenvironments and interactions with immune responses. A comprehensive understanding of these interactions could illuminate novel therapeutic avenues that could enhance the efficacy of glioblastoma treatment protocols.

As research continues, the potential for translational applications derived from this study becomes clearer. Future clinical trials are essential to validate the safety and efficacy of Shikonin in human subjects. If successful, Shikonin could become a cornerstone in novel therapeutic regimens for glioblastoma, ultimately improving patient outcomes.

The science community eagerly anticipates further studies that address the challenges of translating these findings into clinical practice. As this research gains traction, it sets the stage for an exciting period of innovation in glioblastoma therapy, where traditional knowledge intersects with cutting-edge science.

In conclusion, the exploration of Shikonin and its effects on glioblastoma provides not just hope for those affected by this disease, but also emphasizes the shared role of natural products in cancer pharmacology. As researchers delve deeper into the cellular mechanisms associated with EMT, p53, miR-361-5p, and ZEB1, a clearer picture of how to combat glioblastoma will emerge, offering a brighter future for patients and their families.

In light of this research, it is evident that unearthing the complexities of glioblastoma requires a multi-faceted approach. The work by Zhang, Liu, and Wang et al. stands as a pivotal contribution to our understanding, one that may pave the way for future breakthroughs in the fight against one of the most challenging cancers.

Subject of Research: Glioblastoma and the effects of Shikonin on epithelial-mesenchymal transition.

Article Title: Shikonin inhibits epithelial-mesenchymal transition in glioblastoma cells by upregulating p53 and promoting miR-361-5p level to suppress ZEB1 expression.

Article References:

Zhang, F., Liu, Z., Wang, Y. et al. Shikonin inhibits epithelial-mesenchymal transition in glioblastoma cells by upregulating p53 and promoting miR-361-5p level to suppress ZEB1 expression.
BMC Neurosci 26, 37 (2025). https://doi.org/10.1186/s12868-025-00956-6

Image Credits: AI Generated

DOI: 10.1186/s12868-025-00956-6

Keywords: glioblastoma, Shikonin, epithelial-mesenchymal transition, p53, miR-361-5p, ZEB1, cancer therapy.