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Targeting of mutant-p53 and MYC as a novel strategy to inhibit oncogenic SPAG5 activity in triple negative breast cancer.

ABSTRACT: Triple negative breast cancer (TNBC) is an aggressive disease which currently has no effective therapeutic targets and prominent biomarkers. The Sperm Associated antigen 5 (SPAG5) is a mitotic spindle associated protein with oncogenic function in several human cancers. In TNBC, increased SPAG5 expression has been associated with tumor progression, chemoresistance, relapse, and poor clinical outcome. Here we show that high SPAG5 expression in TNBC is regulated by coordinated activity of YAP, mutant p53 and MYC. Depletion of YAP or mutant p53 proteins reduced SPAG5 expression and the recruitment of MYC onto SPAG5 promoter. Targeting of MYC also reduced SPAG5 expression and concomitantly tumorigenicity of TNBC cells. These effects of MYC targeting were synergized with cytotoxic chemotherapy and markedly reduced TNBC oncogenicity in SPAG5-expression dependent manner. These results suggest that mutant p53-MYC-SPAG5 expression can be considered as bona fide predictors of patient's outcome, and reliable biomarkers for effective anticancer therapies.

SUBMITTER: Canu V

PROVIDER: S-EPMC11336084 | biostudies-literature | 2024 Aug

REPOSITORIES: biostudies-literature

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Targeting of mutant-p53 and MYC as a novel strategy to inhibit oncogenic SPAG5 activity in triple negative breast cancer.

Canu Valeria V Vaccarella Sebastiano S Sacconi Andrea A Pulito Claudio C Goeman Frauke F Pallocca Matteo M Rutigliano Daniela D Lev Sima S Strano Sabrina S Blandino Giovanni G

Cell death & disease 20240820 8

Triple negative breast cancer (TNBC) is an aggressive disease which currently has no effective therapeutic targets and prominent biomarkers. The Sperm Associated antigen 5 (SPAG5) is a mitotic spindle associated protein with oncogenic function in several human cancers. In TNBC, increased SPAG5 expression has been associated with tumor progression, chemoresistance, relapse, and poor clinical outcome. Here we show that high SPAG5 expression in TNBC is regulated by coordinated activity of YAP, muta ...[more]

PMID: 39164278

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Project description:We recently established c-Abl as a potent suppressor of triple-negative breast cancer (TNBC) progression through its reactivation of a p53:p21 signaling axis coupled to senescence. Moreover, we observed co-expression of p53 and c-Abl to be essential for normal mammary epithelial cell physiology, as this relationship is lost upon breast cancer progression. Cytoplasmic c-Abl activity is markedly increased in some TNBCs and contributes to disease progression; however, the mechanisms underlying these events remain largely unknown. In addressing this question, we show here that c-Abl is predominantly restricted to the cytoplasm of human MDA-MB-231 TNBC cells, and to the nucleus of human MCF-7 luminal A cells. TTK is a mitotic protein kinase that phosphorylates c-Abl on Thr735, thereby creating a recognition binding motif for 14-3-3 adaptor proteins in response to oxidative stress. By interrogating the METABRIC database, we observed a significant correlation between p53 expression and that of c-Abl and TTK in basal-like breast cancers. Moreover, heterologous expression of TTK in MCF-7 cells significantly stimulated their growth in part via a c-Abl-dependent mechanism. Conversely, depleting TTK expression in MDA-MB-231 cells not only inhibited their organoid growth in 3D-cultures, but also sensitized them to the tumor suppressing activities of c-Abl independent of its subcellular localization. Moreover, we show that mutant p53 forms cytoplasmic complexes with c-Abl, thereby dictating the subcellular localization of c-Abl and the sensitivity of MDA-MB-231 cells to Imatinib. In response to nutrient deprivation, c-Abl:p53 complexes readily accumulate in the nucleus, resulting in the hyperactivation of c-Abl and initiation of its anti-tumor activities. Collectively, we identified a novel mutant p53:c-Abl cytoplasmic signaling complex that promotes MDA-MB-231 cell growth and highlights the contextual cues that confer oncogenic activity to c-Abl in breast cancer.

Project description:BACKGROUND:Triple-negative breast cancer (TNBC) is an aggressive breast cancer subtype that lacks effective therapeutic targets. Sperm-associated antigen 5 (SPAG5) is a mitotic spindle-associated protein that is involved in various biological processes in cervical cancer and bladder urothelial carcinoma. However, the role of SPAG5 in TNBC remains undefined. METHODS:The expression of SPAG5 was examined in TNBC patients via quantitative real-time polymerase chain reaction (qRT-PCR), western blotting, and immunohistochemistry (IHC). The biological functions of SPAG5 in TNBC and the underlying mechanisms were investigated in vitro and in vivo. RESULTS:SPAG5 expression was significantly upregulated in TNBC tissues compared with that in paired adjacent noncancerous tissues (ANTs). High SPAG5 expression was associated with increased lymph node metastasis and high risk of local recurrence. SPAG5 protein expression was significantly associated with poor disease-free survival in TNBC. Gene set enrichment analysis of TNBC data from The Cancer Genome Atlas (TCGA) indicated that high SPAG5 expression was significantly associated with cell cycle and the ATR-BRCA pathway. Functional assays demonstrated that SPAG5 expression promoted tumor growth in vitro and in vivo. In addition, SPAG5-silenced cells were more sensitive to the PARP inhibitor (PARPi) olaparib. Mechanistically, SPAG5 interacted with c-MYC binding protein (MYCBP), thereby increasing MYCBP protein levels and leading to increased c-MYC transcriptional activity, which promoted the expression of the c-MYC target genes: CDC20, CDC25C, BRCA1, BRCA2, and RAD51.Knockdown of MYCBP or c-MYC abolished the SPAG5-induced cell-cycle progression and cell proliferation of TNBC. CONCLUSIONS:Collectively, our results indict that SPAG5 is an efficient prognostic factor in TNBC, and that SPAG5 knockdown increases the sensitivity of TNBC to the PARPi olaparib. SPAG5 promotes tumor growth and DNA repair by increasing c-MYC transcriptional activity via interaction with MYCBP. The SPAG5/MYCBP/c-MYC axis may represent a potential therapeutic target for TNBC treatment.

Project description:Over 80% of triple negative breast cancers express mutant p53. Mutant p53 often gains oncogenic function suggesting that triple negative breast cancers may be driven by p53 protein type. To determine the chromatin targets of this gain-of-function mutant p53 we used inducible knockdown of endogenous gain-of-function mtp53 in MDA-MB-468 cells in conjunction with stable isotope labeling with amino acids in cell culture and subcellular fractionation. We sequenced over 70,000 total peptides for each corresponding reciprocal data set and were able to identify 3010 unique cytoplasmic fraction proteins and 3403 unique chromatin fraction proteins. The present proteomics experiment corroborated our previous experiment-based results that poly ADP-ribose polymerase has a positive association with mutant p53 on the chromatin. Here, for the first time we report that the heterohexomeric minichromosome maintenance complex that participates in DNA replication initiation ranked as a high mutant p53-chromatin associated pathway. Enrichment analysis identified the minichromosome maintenance members 2-7. To validate this mutant p53- poly ADP-ribose polymerase-minichromosome maintenance functional axis, we experimentally depleted R273H mutant p53 and found a large reduction of the amount of minichromosome maintenance complex proteins on the chromatin. Furthermore a mutant p53-minichromosome maintenance 2 direct interaction was detected. Overexpressed mutant p53, but not wild type p53, showed a protein-protein interaction with minichromosome maintenance 2 and minichromosome maintenance 4. To target the mutant p53- poly ADP-ribose polymerase-minichromosome maintenance axis we treated cells with the poly ADP-ribose polymerase inhibitor talazoparib and the alkylating agent temozolomide and detected synergistic activation of apoptosis only in the presence of mutant p53. Furthermore when minichromosome maintenance 2-7 activity was inhibited the synergistic activation of apoptosis was blocked. This mutant p53- poly ADP-ribose polymerase -minichromosome maintenance axis may be useful for theranostics.

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Targeting of mutant-p53 and MYC as a novel strategy to inhibit oncogenic SPAG5 activity in triple negative breast cancer.

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Targeting of mutant-p53 and MYC as a novel strategy to inhibit oncogenic SPAG5 activity in triple negative breast cancer.

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