Identification, validation, and targeting of the mutant p53-PARP-MCM chromatin axis in triple negative breast cancer

Abstract

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.

MCMs in Cancer: Prognostic Potential and Mechanisms

“…MCMs are also involved in apoptosis pathways and inhibit some apoptotic proteins. In triple-negative breast cancer, MCM2-7 complex was reported to participate in a high mutant p53-chromatin-associated pathway [95]. In this research, a protein-protein interaction between overexpressed mutant p53 and MCM2/4 was directly detected.…”

“…Based on public databases, strong positive coexpression was observed among MCM2-7 genes [94]. In triple-negative breast cancer, protein-protein interaction between Mcm2 and Mcm4 was verified with proximity ligation assay [95].…”

The Signaling Duo CXCL12 and CXCR4: Chemokine Fuel for Breast Cancer Tumorigenesis

“…Specifically, CXCL12 seems to play a detrimental role in breast cancer; its expression in patients has been extensively studied. High CXCL12 levels are present in patients with lymph node and brain metastasis and inversely correlate with the overall survival [ 32 , 33 , 34 ]. CXCL12 in breast cancer is typically released from stromal cells, and in TNBC it is secreted by a specific immunosuppressive subset of fibroblasts [ 33 , 35 ].…”

“…They are stronger inducers of cancer cell migration. High levels of CXCL12 and CXCL16 in CAFs from human brain cancer metastasis attract breast cancer cells via the CXCR4-CXCL12 and CXCR6-CXCL16 pathways [ 33 ].…”

Update on the Biomarkers for Targeted Therapy in Triple-Negative Breast Cancer.

“…It was functionally demonstrated that miR-498 inhibits BRCA1 in breast cancer cell lines, and showed that inhibition of miR-498 led to reduced proliferation in the triple negative cell line Hs578T and results indicate that miR-498 regulates BRCA1 expression in breast cancer and its overexpression could contribute to the pathogenesis of sporadic TNBC via BRCA1 downregulation. 21 an oncomiR was proven to regulate PTEN by targeting the mRNA 3′UTR, leading to an anti-apoptosis effect in TNBC (32). Specifically, miR-21 exhibits a higher expression level in TNBC than non-TNBC, and it is positively correlated with a poor clinical outcome.…”