Valproic Acid Regulates HR and Cell Cycle Through MUS81-pRPA2 Pathway in Response to Hydroxyurea

Abstract

Breast cancer is the primary problem threatening women’s health. The combined application of valproic acid (VPA) and hydroxyurea (HU) has a synergistic effect on killing breast cancer cells, but the molecular mechanism remains elusive. Replication protein A2 phosphorylation (pRPA2), is essential for homologous recombination (HR) repair and cell cycle. Here we showed that in response to HU, the VPA significantly decreased the tumor cells survival, and promoted S-phase slippage, which was associated with the decrease of pCHK1 and WEE1/pCDK1-mediated checkpoint kinases phosphorylation pathway and inhibited pRPA2/Rad51-mediated HR repair pathway; the mutation of pRPA2 significantly diminished the above effect, indicating that VPA-caused HU sensitization was pRPA2 dependent. It was further found that VPA and HU combination treatment also resulted in the decrease of endonuclease MUS81. After MUS81 elimination, not only the level of pRPA2 was abolished in response to HU treatment, but also VPA-caused HU sensitization was significantly down-regulated through pRPA2-mediated checkpoint kinases phosphorylation and HR repair pathways. In addition, the VPA altered the tumor microenvironment and reduced tumor burden by recruiting macrophages to tumor sites; the Kaplan-Meier analysis showed that patients with high pRPA2 expression had significantly worse survival. Overall, our findings demonstrated that VPA influences HR repair and cell cycle through down-regulating MUS81-pRPA2 pathway in response to HU treatment.

VPA mediates bidirectional regulation of cell cycle progression through the PPP2R2A-Chk1 signaling axis in response to HU

“…We previously demonstrated that VPA inhibits cancer cell growth after HU treatment [ 10 , 32 ]. In the current study, we found that normal 16HBE (Fig.…”

“…We next used a primary rat model of DMBA-induced breast cancer that we had described previously [ 10 , 32 ] to triangulate the in vitro data. In this animal model, we concurred with previous studies that VPA + HU treatment produced a significant reduction in tumor volume (Fig.…”

“…Hydroxyurea (HU) depletes dNTPs to induce stalled or collapsed replication forks and is a commonly used antitumor drug with a known toxicology profile [ 7 , 8 ]. Our previous study showed that tumor cells treated with 2 mM HU for 18 h resulted in one-end DNA double-strand breaks, activated the checkpoint kinase-1 (Chk1) signaling pathway and arrested tumor cells in the S phase [ 9 , 10 ]. Chk1 coordinates the response to DNA damage [ 11 – 13 ] and is activated through its phosphorylation by the ataxia telangiectasia and RAD3-related (ATR) protein kinase [ 10 , 14 ].…”

“…Our previous study showed that tumor cells treated with 2 mM HU for 18 h resulted in one-end DNA double-strand breaks, activated the checkpoint kinase-1 (Chk1) signaling pathway and arrested tumor cells in the S phase [ 9 , 10 ]. Chk1 coordinates the response to DNA damage [ 11 – 13 ] and is activated through its phosphorylation by the ataxia telangiectasia and RAD3-related (ATR) protein kinase [ 10 , 14 ]. Chk1 signaling promotes cell cycle arrest and prevents the entry of cells with damaged or incompletely replicated DNA into mitosis, especially in cells lacking checkpoints in the cell cycle G1 phase [ 13 , 15 , 16 ].…”

“…Some HDACis are approved in cancer treatment [ 28 – 31 ]. In our previous study, VPA, a class I and II HDACi, has been shown to increase the sensitivity of tumor cells to HU treatment; for example, VPA reduced S-phase arrest of the cell cycle and thus has a chemo-sensitizing role in response to HU-induced replication arrest by inhibiting Chk1 signaling [ 10 , 32 , 33 ]. The precise molecular mechanisms by which VPA sensitizes tumor cells through regulating checkpoint kinases in the various stages of the cell cycle and whether VPA affords protection on normal and abscopal cells to avoid HU toxicity remains elusive.…”