Coactivator-associated arginine methyltransferase 1 (CARM1), identified 20 years ago as a coregulator of transcription, is an enzyme that catalyzes arginine methylation of proteins. Beyond its ...well-established involvement in the regulation of transcription, the physiological functions of CARM1 are still poorly understood. However, recent studies have revealed novel roles of CARM1 in autophagy, metabolism, paraspeckles, and early development. In addition, CARM1 is emerging as an attractive therapeutic target and a drug response biomarker for certain types of cancer. Here, we provide a comprehensive overview of the structure of CARM1 and its post-translational modifications, its various functions, apart from transcriptional coactivation, and its involvement in cancer.
In addition to the initially described activity as a transcriptional coactivator, the emerging functions of CARM1 include autophagy, metabolism, early development, pre-mRNA splicing and export, and localization to paraspeckles.Aside from post-translationally modifying its substrates, CARM1 itself undergoes several modifications, such as phosphorylation, O-GlcNAcylation, ubiquitination, and auto-methylation.CARM1 is composed of a central catalytic and unique N- and C-terminal domains that are important for binding to its partners.CARM1 is the only known PRMT to preferentially methylate a proline-rich sequence.A spliced isoform of CARM1 is the major variant expressed in BCs.CARM1 has now emerged as an attractive therapeutic target/biomarker of response for various types of cancer.Specific CARM1 inhibitors show antitumor activity in vivo in myeloma models.
Triple negative breast cancer (TNBC) is an aggressive breast cancer subtype. Non-available targeted therapy for TNBC represents its biggest treatment challenge. Thus, finding new promising effective ...drugs is urgently needed. In the present study, we investigated how berberine, a natural isoquinoline, impairs the survival of TNBC cells in both cellular and molecular levels. Our experimental model was based on the use of eight TNBC cell lines: MDA-MB-468, MDA-MB-231, HCC70, HCC38, HCC1937, HCC1143, BT-20, and BT-549. Berberine was cytotoxic against all treated TNBC cell lines. The most sensitive cell lines were HCC70 (IC
= 0.19 µM), BT-20 (IC
= 0.23 µM) and MDA-MB-468 (IC
= 0.48 µM). Using flow cytometry techniques, berberine, at 0.5 and 1 µM for 120 and 144 h, not only induced cell cycle arrest, at G1 and/or G2/M phases, but it also triggered significant apoptosis. At the molecular level, these results are consistent with the expression of their related proteins using Western blot assays. Interestingly, while berberine was cytotoxic against TNBC cells, it had no effect on the viability of normal human breast cells MCF10A cultured in a 3D matrigel model. These results suggest that berberine may be a good potential candidate for TNBC drug development.
The triple-negative breast cancer (TN BC) subtype is the most aggressive form of invasive BC. Despite intensive efforts to improve BC treatments, patients with TN BC continue to exhibit poor ...survival, with half developing resistance to chemotherapy. Here we identify autophagy as a key mechanism in the progression and chemoresistance of a subset of TN tumors. We demonstrate that LC3B, a protein involved in autophagosome formation, is a reliable marker of poor prognosis in TN BC, validating this prognostic value at both the mRNA and protein levels in several independent cohorts. We also show that LC3B has no prognostic value for other BC subtypes (Luminal or HER2 BC), thus revealing a specific impact of autophagy on TN tumors. Autophagy is essential for the proliferative and invasive properties in 3D of TN BC cells characterized by high LC3B levels. Interestingly, the activity of the transcriptional co-activator YAP1 (Yes-associated protein 1) is regulated by the autophagy process and we identify YAP1 as a new actor in the autophagy-dependent proliferative and invasive properties of high-LC3B TN BC. Finally, inhibiting autophagy by silencing ATG5 or ATG7 significantly impaired high-LC3B TN tumor growth in vivo. Moreover, using a patient-derived TN tumor transplanted into mice, we show that an autophagy inhibitor, chloroquine, potentiates the effects of chemotherapeutic agents. Overall, our data identify LC3B as a new prognostic marker for TN BC and the inhibition of autophagy as a promising therapeutic strategy for TN BC patients.
The PI3K/AKT pathway is considered to play a major role in bladder carcinogenesis, but its relationships with other molecular alterations observed in bladder cancer remain unknown. We investigated ...PI3K/AKT pathway activation in a series of human bladder urothelial carcinomas (UC) according to PTEN expression, PTEN deletions and FGFR3, PIK3CA, KRAS, HRAS, NRAS and TP53 gene mutations. The series included 6 normal bladder urothelial samples and 129 UC (Ta n = 25, T1 n = 34, T2–T3–T4 n = 70). Expression of phospho‐AKT (pAKT), phospho‐S6‐Ribosomal Protein (pS6) (one downstream effector of PI3K/AKT pathway) and PTEN was evaluated by reverse phase protein Array. Expression of miR‐21, miR‐19a and miR‐222, known to regulate PTEN expression, was also evaluated. pAKT expression levels were higher in tumors than in normal urothelium (p < 0.01), regardless of stage and showed a weak and positive correlation with pS6 (Spearman coefficient RS = 0.26; p = 0.002). No association was observed between pAKT or pS6 expression and the gene mutations studied. PTEN expression was decreased in PTEN‐deleted tumors, and in T1 (p = 0.0089) and T2–T3–T4 (p < 0.001) tumors compared to Ta tumors; it was also negatively correlated with miR‐19a (RS = −0.50; p = 0.0088) and miR‐222 (RS = −0.48; p = 0.0132), but not miR‐21 (RS = −0.27; p = 0.18) expression. pAKT and PTEN expressions were not negatively correlated, and, on the opposite, a positive and moderate correlation was observed in Ta (RS = 0.54; p = 0.0056) and T1 (RS = 0.56; p = 0.0006) tumors. Our study suggests that PI3K/AKT pathway activation occurs in the entire spectrum of bladder UC regardless of stage or known most frequent molecular alterations, and independently of low PTEN expression.
What's new?
Although a suspected player in bladder carcinogenesis, involvement of the PI3K/AKT pathway in human bladder urothelial carcinoma (UC) has been little explored. Here, reverse phase protein array was used to investigate PI3K/AKT pathway activation and expression of PTEN, a PI3K/AKT pathway downregulator, in a series of human UCs characterized by genetic alterations frequently observed in UC. Analysis revealed elevated pAKT expression and PI3K/AKT pathway activation across UCs, despite the different molecular alterations and independent of the loss of PTEN. The findings support the idea that the PI3K/AKT pathway is of therapeutic significance in UC.
Triple-negative breast cancer (TNBC) represents a subgroup of breast cancers (BC) associated with the most aggressive clinical behavior. No targeted therapy is currently available for the treatment ...of patients with TNBC. In order to discover potential therapeutic targets, we searched for protein kinases that are overexpressed in human TNBC biopsies and whose silencing in TNBC cell lines causes cell death. A cohort including human BC biopsies obtained at Institut Curie as well as normal tissues has been analyzed at a gene-expression level. The data revealed that the human protein kinase monopolar spindle 1 (hMPS1), also known as TTK and involved in mitotic checkpoint, is specifically overexpressed in TNBC, compared to the other BC subgroups and healthy tissues. We confirmed by immunohistochemistry and reverse phase protein array that TNBC expressed higher levels of TTK protein compared to the other BC subgroups. We then determined the biological effects of TTK depletion by RNA interference, through analyses of tumorigenic capacity and cell viability in different human TNBC cell lines. We found that RNAi-mediated depletion of TTK in various TNBC cell lines severely compromised their viability and their ability to form colonies in an anchorage-independent manner. Moreover, we observed that TTK silencing led to an increase in H2AX phosphorylation, activation of caspases 3/7, sub-G1 cell population accumulation and high annexin V staining, as well as to a decrease in G1 phase cell population and an increased aneuploidy. Altogether, these data indicate that TTK depletion in TNBC cells induces apoptosis. These results point out TTK as a protein kinase overexpressed in TNBC that may represent an attractive therapeutic target specifically for this poor prognosis associated subgroup of breast cancer.
The canonical Wnt/β-catenin pathway is activated in triple-negative breast cancer (TNBC). The activation of this pathway leads to the expression of specific target genes depending on the cell/tissue ...context. Here, we analyzed the transcriptome of two different TNBC cell lines to define a comprehensive list of Wnt target genes. The treatment of cells with Wnt3a for 6h up-regulated the expression (fold change > 1.3) of 59 genes in MDA-MB-468 cells and 241 genes in HCC38 cells. Thirty genes were common to both cell lines. Beta-catenin may also be a transcriptional repressor and we found that 18 and 166 genes were down-regulated in response to Wnt3a treatment for 6h in MDA-MB-468 and HCC38 cells, respectively, of which six were common to both cell lines. Only half of the activated and the repressed transcripts have been previously described as Wnt target genes. Therefore, our study reveals 137 novel genes that may be positively regulated by Wnt3a and 104 novel genes that may be negatively regulated by Wnt3a. These genes are involved in the Wnt pathway itself, and also in TGFβ, p53 and Hedgehog pathways. Thorough characterization of these novel potential Wnt target genes may reveal new regulators of the canonical Wnt pathway. The comparison of our list of Wnt target genes with those published in other cellular contexts confirms the notion that Wnt target genes are tissue-, cell line- and treatment-specific. Genes up-regulated in Wnt3a-stimulated cell lines were more strongly expressed in TNBC than in luminal A breast cancer samples. These genes were also overexpressed, but to a much lesser extent, in HER2+ and luminal B tumors. We identified 72 Wnt target genes higher expressed in TNBCs (17 with a fold change >1.3) which may reflect the chronic activation of the canonical Wnt pathway that occurs in TNBC tumors.
Between 30% and 70% of patients with breast cancer have pre-existing chronic conditions, and more than half are on long-term non-cancer medication at the time of diagnosis. Preliminary ...epidemiological evidence suggests that some non-cancer medications may affect breast cancer risk, recurrence, and survival. In this nationwide cohort study, we assessed the association between medication use at breast cancer diagnosis and survival. We included 235,368 French women with newly diagnosed non-metastatic breast cancer. In analyzes of 288 medications, we identified eight medications positively associated with either overall survival or disease-free survival: rabeprazole, alverine, atenolol, simvastatin, rosuvastatin, estriol (vaginal or transmucosal), nomegestrol, and hypromellose; and eight medications negatively associated with overall survival or disease-free survival: ferrous fumarate, prednisolone, carbimazole, pristinamycin, oxazepam, alprazolam, hydroxyzine, and mianserin. Full results are available online from an interactive platform ( https://adrenaline.curie.fr ). This resource provides hypotheses for drugs that may naturally influence breast cancer evolution.
Breast cancer is composed of distinct subgroups, triple-negative breast cancer (TNBC), human epidermal growth factor receptor-2 (HER2), luminal A, and luminal B, which are associated with different ...prognosis. MEP50 is the main partner of the arginine methyltransferase PRMT5 required for its enzymatic activity. Here, we examined MEP50 expression in the different breast cancer subgroups from the transcriptomic data obtained on human breast cancer samples and on normal breast tissues in two cohorts (Curie, n = 141; The Cancer Genome Atlas—TCGA, n = 788). We observed higher levels of MEP50 mRNA in TNBC (Curie, n = 41; TCGA, n = 106) compared to the other breast cancer subgroups and normal breast tissues. Using an online KM-plotter database, which allows survival analyses in a larger number of breast cancer patients, we found that high MEP50 mRNA levels were associated with a more favorable recurrence-free survival (RFS) in TNBC (n = 953, p = 1.2 × 10−4) and luminal B (n = 1353, p = 0.013) tumors, whereas high PRMT5 mRNA levels were associated with worse RFS in these two subgroups (TNBC: n = 442, p = 1.0 × 10−4; luminal B: n = 566, p = 6.8 × 10−3). We next determined the expression and the subcellular localization of MEP50 protein by immunohistochemistry (IHC) in our Curie cohort of breast cancer (n = 94) and normal tissues (n = 7) using a validated MEP50 antibody. MEP50 was more expressed in breast tumors compared to normal breast tissues (p = 0.02). MEP50 was more localized to the cytosol in breast cancer cells compared to normal breast tissue (p = 4 × 10−4), and was more found at the plasma membrane in normal tissues compared to breast tumors (p = 0.01). We also evaluated PRMT5 activity by IHC in our Curie cohort using a validated antibody (H4R3me2s) detecting histone H4 symmetrically dimethylated on Arg3. High levels of H4R3me2s were found in normal breast tissues, whereas the lowest levels of H4R3me2s were observed in TNBC and HER2 breast cancer subgroups. Altogether, our study reports the expression of the PRMT5 cofactor (MEP50) and substrate (H4R3me2s) in breast cancer and highlights the association of PRMT5 and MEP50 mRNA with prognosis in luminal B and TNBC breast cancer subgroups and certain TNBC subtypes.
Therapeutic strategies targeting Homologous Recombination Deficiency (HRD) in breast cancer requires patient stratification. The LST (Large‐scale State Transitions) genomic signature previously ...validated for triple‐negative breast carcinomas (TNBC) was evaluated as biomarker of HRD in luminal (hormone receptor positive) and HER2‐overexpressing (HER2+) tumors. The LST genomic signature related to the number of large‐scale chromosomal breakpoints in SNP‐array tumor profile was applied to identify HRD in in‐house and TCGA sets of breast tumors, in which the status of BRCA1/2 and other genes was also investigated. In the in‐house dataset, HRD was predicted in 5% (20/385) of sporadic tumors luminal or HER2+ by the LST genomic signature and the inactivation of BRCA1, BRCA2 or RAD51C confirmed this prediction in 75% (12/16) of the tested cases. In 14% (6/43) of tumors occurring in BRCA1/2 mutant carriers, the corresponding wild‐type allele was retained emphasizing the importance of determining the tumor status. In the TCGA luminal and HER2+ subtypes HRD incidence was estimated at 5% (18/329, 95%CI: 5–8%) and 2% (1/59, 95%CI: 2–9%), respectively. In TNBC cisplatin‐based neo‐adjuvant clinical trials, HRD is shown to be a necessary condition for cisplatin sensitivity. This analysis demonstrates the high performance of the LST genomic signature for HRD detection in breast cancers, which suggests its potential as a biomarker for genetic testing and patient stratification for clinical trials evaluating platinum salts and PARP inhibitors.
What's new?
The BRCA1/2 tumor suppressors play a key role in DNA repair by homologous recombination, and inactivation of these factors is associated with high breast and ovarian cancer risk. Here, the authors performed a comprehensive study of breast tumors screening for homologous recombination deficiency (HRD) measured by large‐scale chromosomal breaks. They find good overlap between HRD‐positivity and BRCA mutations, but also identify a small subset of HRD+ tumors in which the sole determination of BRCA1/2 mutational status is insufficient. This points to HRD determination as a potentially important method to find patients benefiting most from emerging new therapies such as PARP inhibitors.
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