Expression of Cyclin D1 protein in residual tumor after neoadjuvant chemotherapy for breast cancer.

PURPOSE: Hormone receptor (HR)-positive breast cancer (BC) shows a poor response to neoadjuvant chemotherapy (NACT). New treatment targets like the Cyclin D1-CDK4/CDK6 complex are promising adjuvant/post-neoadjuvant therapeutic strategies. Evaluating Cyclin D1 overexpression in residual tumor could recognize those patients that benefit most from such post-neoadjuvant treatment. In this study, we determined Cyclin D1 expression in residual BC after NACT. Secondary aims were to correlate Cyclin D1 expression levels with clinicopathological parameters and to assess its prognostic value after NACT. METHODS: We retrospectively assessed the nuclear expression of Cyclin D1 on tissue microarrays with residual tumor from 284 patients treated in the neoadjuvant GeparTrio (n = 186) and GeparQuattro (n = 98) trials. Evaluation was performed with a standardized immunoreactive score (IRS) after selecting a cut-off value. RESULTS: A high expression level (IRS ≥ 6) of Cyclin D1 was found in 37.3% of the assessed specimens. An increased Cyclin D1 expression was observed in HR-positive tumors, compared to HR-negative tumors (p = 0.02). Low Cyclin D1 levels correlated with clinical tumor stage 1-3 (p = 0.03). Among patients with HR-positive/Her2-negative tumors and high Cyclin D1 expression, a better disease-free survival (DFS) was graphically suggested, but not significant (p = 0.21). CONCLUSION: Our study demonstrates a measurable nuclear expression of Cyclin D1 in post-neoadjuvant residual tumor tissue of HR-positive BC. Cyclin D1 expression was not prognostic for DFS after NACT. Our results and defined cut-off suggest that the marker can be used to stratify tumors according to protein expression levels. Based on this, a prospective evaluation is currently performed in the ongoing Penelope-B trial.

Reduction of HIF-1α/PD-L1 by Catalytic Topoisomerase Inhibitor Induces Cell Death Through Caspase Activation in Cancer Cells Under Hypoxia.

BACKGROUND/AIM: Under severe hypoxia, cellular apoptosis is induced through hypoxia-inducible factor 1, alpha subunit (HIF-1α)-dependent P53 accumulation and P53 phosphorylation via ataxia telangiectasia mutated and ataxia telangiectasia and RAD3-related (ATR) activation via replication stress-induced DNA damage response (DDR) activation. We previously demonstrated that the topoisomerase I catalytic inhibitor, 3-O-(2'E,4'Z-decadienoyl)-20-O-acetylingenol (3EZ,20Ac-ingenol), induced apoptosis in Jeko-1 and Panc-1 cells, both of which show cyclin D1 overexpression. After progression to the S phase facilitated by nuclear cyclin D1, an intra S phase checkpoint was induced in the presence of 3EZ,20Ac-ingenol, by ATR activation in response to replication stress-induced DDR. MATERIALS AND METHODS: In this study, we examined whether 3-O-(2'E,4'E-decadienoyl)-20-O-acetylingenol (3EE,20Ac-ingenol) might induce a higher degree of P53 phosphorylation and additional HIF-1α and P53 accumulation in response to replication stress-induced DDR activation under hypoxic conditions than under normoxic conditions, by controlling ATR activation. RESULTS: In the Panc-1 cells, 3EE,20Ac-ingenol induced P53 activation and HIF-1α-dependent P53 accumulation through cooperative ATR activation via hypoxia-induced DDR activation. Jeko-1 cells showed slight HIF-1α accumulation under hypoxia, but HIF-1α-dependent 53 accumulation was not observed in the presence of 3EE,20Ac-ingenol, so that the cells remained resistant to hypoxia. CONCLUSION: 3EE,20Ac-ingenol induces an intricate interplay between P53 and HIF-1α accumulation via ATR activation that results in a high P53 accumulation, which promoted transient expression and early disappearance of HIF-1α, accelerating cell death. Strong P53 accumulation and consequent phosphatase and tensin homolog deleted on chromosome 10 activation in Panc-1 cells also reduced HIF-1α accumulation and programmed death-ligand 1 expression, which resulted in intense apoptosis.

3EZ, 20Ac-ingenol induces cell-specific apoptosis in cyclin D1 over-expression through the activation of ATR and downregulation of p-Akt.

Acute lymphoblastic leukemia (ALL) samples exhibit an activated PI3K/Akt pathway, which suggests a general role of Akt in the development of leukemia. We have previously used western blot analysis to show that the catalytic topoisomerase (topo) inhibitor, 3EZ, 20Ac-ingenol, induced DNA damage response (DDR), which activated ATR, downregulated p-Akt through upregulation of PTEN level, and led to cell cycle arrest or apoptosis. In this study, we used ATR or PTEN siRNA and observed that the specific cell arrest and apoptosis of BALL-1 cells in DDR caused by 3EZ, 20Ac-ingenol was dependant on activation of ATR and downregulation of nuclear p-Akt through upregulation of PTEN. Moreover, some B cell lymphomas among ALLs overexpress cyclin D1. The DDR induced during the S-phase with 3EZ, 20Ac-ingenol treatment was increased by the intra S-phase checkpoint response that was triggered by the loss of nuclear cyclin D1 regulation in BALL-1 cells overexpressing cyclin D1. Although topo 1 catalytic inhibitors induce a decatenation checkpoint and subsequent G2/M phase arrest, the decatenation checkpoint caused by 3EZ, 20Ac-ingenol induced apoptosis only in the BALL-1 cells that accumulated cyclin D1.