It is suggested that PRMT5 inhibition by CMP5 is associated with reduced TNBC cell viability, impaired migration, increased expression of apoptosis-associated regulators and enhanced apoptotic cell death as measured by Annexin V/PI analysis in vitro.
Abstract
Background: PRMT5, or protein arginine methyltransferase 5, is recognized as an epigenetic regulator that suppresses gene transcription through symmetric dimethylation of histone arginine residues, including histone H4 arginine 3 (H4R3me2s) and histone H3 arginine 8 (H3R8me2s), modifications associated with chromatin condensation and transcriptional repression. PRMT5-mediated methylation has been associated with recruitment of polycomb repressive complex 2 (PRC2) and enhancer of zeste homolog 2 (EZH2)-mediated H3K27me3 deposition, contributing to stable repression of tumor suppressor genes and apoptosis-related effectors in breast cancer. Methods: The molecular and functional impacts of PRMT5 inhibition were studied in TNBC cell lines with a pharmacological inhibitor (CMP5). Cellular responses were evaluated using a viability assay, qPCR, Western blotting, Annexin V/PI staining, and transwell migration/proliferation assays. Results: PRMT5 inhibition substantially reduced TNBC viability in a time- and dose-dependent manner. EZH2 was downregulated, whereas the tumor suppressor retinoblastoma-like protein 2 (RBL2) was induced, concomitant with low expression of Cyclin D1. These changes were accompanied by upregulation of pro-apoptotic effectors (Caspase-3, Caspase-10, death-associated protein 1 (DAP1), and BCL2-associated x protein (BAX) and repression of the pro-survival B-cell lymphoma 2 (BCL2), consistent with apoptosis-associated molecular responses. Functionally, CMP5 treatment was associated with reduced migratory behavior in TNBC cells under the experimental conditions tested. Conclusions: These findings suggest that PRMT5 inhibition by CMP5 is associated with reduced TNBC cell viability, impaired migration, increased expression of apoptosis-associated regulators and enhanced apoptotic cell death as measured by Annexin V/PI analysis in vitro. Further mechanistic and in vivo studies are required to clarify the therapeutic relevance of PRMT5 inhibition in TNBC.
Mounting evidence has shown that histone acetyltransferase binding to ORC1 (HBO1) serves as an oncoprotein, warranting the use of the small molecule inhibitor WM-3835 for cancer therapy. However, HBO1 is ubiquitously expressed in both tumor and normal tissues, with potential to increase the risk of systemic toxicity. This unmet need highlights the importance of identifying suitable biomarkers to predict the sensitivity to HBO1 inhibitor. Here, we show that ATR, a key regulator of DNA replication stress, is a novel interacting partner of HBO1. Additionally, we reveal a regulatory function of HBO1 in DNA replication stress responses, in an ATR-dependent manner. Mechanistically, ATR mediated HBO1 Ser50/53 phosphorylation interferes with the genomic binding of HBO1 and regulates gene expression. Notably, overexpression of HBO1 mutated at the ATR phosphorylation site (S50/53A) dampens the expression of DNA repair related genes and suppresses tumor colony formation, consistent with the observations of WM-3835 treatment. Inhibition of ATR significantly antagonized the sensitivity to WM-3835 treatment. Collectively, our findings uncovered a previously unidentified role of HBO1 in the regulation of replication stress and discovered ATR as a potential biomarker for WM-3835 treatment.
C. Zong, Jiamin Zhang, Zhe Zhang et al.· Journal of Biological Chemis...· 0 citations
Abstract Background Doxorubicin, an anthracycline chemotherapeutic agent, is widely used in diffuse large B‑cell lymphoma (DLBCL) treatment, yet its clinical efficacy is often compromised by drug resistance. Protein arginine methyltransferase 7 (PRMT7) is a methyltransferase implicated in tumourigenesis and cancer progression. However, its precise role and underlying mechanisms in DLBCL progression and doxorubicin resistance remain unclear. Methods We analysed PRMT7 expression in DLBCL cell lines and patient specimens using bioinformatic databases, western blotting, reverse Transcription Polymerase Chain Reactionand immunohistochemistry. Functional studies were performed in DLBCL cell lines through CRISPR/Cas9‑mediated knockout and overexpression systems, combined with in vitro assays for proliferation, apoptosis and doxorubicin sensitivity, as well as in vivo xenograft models. Mechanistically, co‑immunoprecipitation, arginine methylation assays and immunofluorescence were employed to characterise the PRMT7–Forkhead box K (FOXK)1/2–Dishevelled Segment Polarity Protein 2 (DVL2) axis and its modulation of Wnt/ (beta) β‑catenin signalling. In addition, we designed and evaluated FOXK‑methylation‑competitive inhibitory peptides for their capacity to sensitise DLBCL cells to doxorubicin. Results Our study revealed that PRMT7 is upregulated in DLBCL and promotes both tumour proliferation and doxorubicin resistance. Conversely, knockdown of PRMT7 inhibited DLBCL cell proliferation and enhanced sensitivity to doxorubicin. Mechanistically, PRMT7 catalyses arginine methylation of FOXK1 at arginine (R) 191 and FOXK2 at R144, which markedly increases their binding affinity for DVL2 and facilitates DVL2 nuclear translocation. This event leads to constitutive activation of the Wnt/β‑catenin signalling pathway. Importantly, we developed a FOXK‑derived peptide that competitively inhibits FOXK methylation, suppresses Wnt/β‑catenin signalling and significantly potentiates the antitumour efficacy of doxorubicin in DLBCL. Conclusion Our findings indicate that the PRMT7/FOXK/DVL2/Wnt‐β‐catenin axis serves as a novel driver of DLBCL progression and doxorubicin resistance. Targeting the PRMT7–FOXK methylation interface may offer a potential therapeutic approach for overcoming doxorubicin resistance in DLBCL, although further validation in clinical settings is warranted. Key points Upregulated PRMT7 drives DLBCL progression and doxorubicin resistance PRMT7 methylates FOXK1 at arginine 191 and FOXK2 at arginine 144 FOXK1/2 methylation enhances interaction with DVL2 and promotes nuclear translocation FOXK1/2 methylation dactivate Wnt/ß-catenin signaling through DVL2 nuclear entry A novel peptide inhibitory blocks FOXK1/2 methylation and restores doxorubicin sensitivity
Xiao-Yan Zhao, J. Zhang, Yao Teng et al.· Clinical and Translational M...· 0 citations
Chemoresistance remains a major barrier in colorectal cancer (CRC) therapy. Through epigenetic compound screening in patient-derived organoids (PDOs), we identified CX6258.HCl as a potent growth inhibitor. Treatment with CX6258.HCl significantly inhibited cell mitosis and induced apoptosis in CRC cell lines. Mechanistically, CX6258.HCl binds the D687 residue within HASPIN's kinase domain, suppressing H3T3 phosphorylation (H3T3ph). This triggers an epigenetic cascade: loss of H3T3ph upregulates demethylase KDM5B pre-mRNA, depleting H3K4me3 at promoters of Kinesin family member (KIFC1/KIF10/KIF14). Consequently, microtubule dynamics are disrupted, leading to mitotic arrest. Target specificity was validated genetically via HASPIN-D687A mutation. In vivo, CX6258.HCl suppressed CRC xenograft growth and further enhanced 5-FU-mediated tumor suppression without obvious histological injury in major organs. Clinically, elevated H3T3ph levels in human CRC tissues were associated with Ki67-positive proliferative tumor regions, suggesting that H3T3ph may represent a proliferation-associated marker in CRC. Together, our findings identify the HASPIN/H3T3ph-KDM5B-H3K4me3-KIF axis as a targetable antimitotic pathway and support therapeutic inhibition of HASPIN/H3T3ph as a potential strategy for CRC.
Tong Wu, Yaguang Zhang, Sicheng Liu et al.· Molecular Cancer· 0 citations
Oxidized low density lipoprotein receptor 1 (OLR1) regulates glycolytic metabolism, lactate production and carcinogenesis, but its role in papillary thyroid cancer (PTC) is still unclear. The present study aimed to investigate the effect of OLR1 modification on PTC cellular functions and its interaction with histone H3 lysine 18 lactylation (H3K18la) and downstream molecular mechanisms. Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1; OLR1 coding protein) and H3K18la expression was detected in twenty pairs of PTC and non-tumor tissues by immunohistochemistry. Overexpressed OLR1 and small interfering (si)RNA were transfected into two PTC cell lines (TPC-1 and IHH-4 cells). After which, oxamate (to reduce lactate production) or LEM domain containing 1 (LEMD1) siRNA combined with OLR1 overexpression plasmid was used to treat PTC cell lines. Furthermore, PTC xenograft mice were constructed for in vivo verification. LOX-1 and H3K18la expression was upregulated in PTC tissues versus non-tumor tissues and LOX-1 expression positively associated with H3K18la expression in PTC tissues. OLR1 overexpression facilitated cell proliferation and invasion but repressed cell apoptosis in PTC cell lines, while OLR1 siRNA revealed an opposite effect. OLR1 positively regulated lactate production, H3K18la expression and LEMD1 expression. Furthermore, a chromatin immunoprecipitation assay confirmed a direct binding between H3K18la and the LEMD1 promoter. H3K18la expression was lowered by oxamate-induced lactate reduction, which also attenuated the effect of OLR1 overexpression on PTC cell proliferation, apoptosis and invasion and downregulated LEMD1 expression. Moreover, LEMD1 siRNA weakened the effect of OLR1 overexpression on PTC cell proliferation, apoptosis and invasion. In vivo experiments revealed that OLR1 overexpression elevated tumor volume and weight and upregulated the expressions of LEMD1 and H3K18la; while LEMD1 knockdown reduced tumor volume and weight. In conclusion, OLR1 promotes PTC proliferation and invasion, while represses apoptosis through regulating H3K18la-mediated LEMD1.
Jia-wei Yu, Rui Pang, Bo Liu et al.· Oncology Report· 0 citations
