TOP2A Protein Research Articles

Expression and biological significance of topoisomerase II α (TOP2A) in oral squamous cell carcinoma

BackgroundTopoisomerase II α(TOP2A) is usually highly expressed in rapidly proliferating cells, and its expression is regulated by cell cycle. The relationship between TOP2A and oral squamous cell carcinoma (OSCC) needs further study.MethodsTOP2A immunoreactivity was analyzed using immunohistochemical (IHC) staining analysis in specimens from 20 OSCC patients. Based on the high-throughput sequencing and gene microarray database, the expression of TOP2A mRNA in OSCC was calculated and its ability to identify OSCC tissues was evaluated by diagnostic analysis. CRISPR screen was used to select the genes necessary for OSCC cell growth, and the gene set was analyzed for function enrichment. Single-cell RNA sequencing analysis was conducted to evaluate the expression level of TOP2A mRNA in OSCC cells.ResultsCompared with normal oral tissues, the expression of TOP2A protein was up-regulated in OSCC tissues. A total of 1240 OSCC and 428 non-OSCC oral tissue samples were included based on high-throughput dataset retrieval, and it was confirmed that TOP2A mRNA was highly expressed in OSCC tissues [SMD = 1.51 (95% CI 0.94–2.07), sROC AUC = 0.96 (95% CI 0.94–0.98)]. As an essential gene for OSCC cell growth, single-cell RNA sequencing data also confirmed that TOP2A mRNA expression was up-regulated in OSCC cells.ConclusionThe up-regulation of TOP2A may play a pivotal role in OSCC.

SUMO Proteomics Analyses Identify Protein Inhibitor of Activated STAT-Mediated Regulatory Networks Involved in Cell Cycle and Cell Proliferation.

Protein inhibitor of activated STAT (PIAS) proteins are E3 SUMO ligases playing important roles in protein stability and signaling transduction pathways. PIAS proteins are overexpressed in the triple-negative breast cancer cell line MDA-MB-231, and PIAS knockout (KO) results in a reduction in cell proliferation and cell arrest in the S phase. However, the molecular mechanisms underlying PIAS functions in cell proliferation and cell cycle remain largely unknown. Here, we used quantitative SUMO proteomics to explore the regulatory role of PIAS SUMO E3 ligases upon CRISPR/Cas9 KO of individual PIAS. A total of 1422 sites were identified, and around 10% of SUMO sites were regulated following KO of one or more PIAS genes. We identified protein substrates that were either specific to individual PIAS ligase or regulated by several PIAS ligases. Ki-67 and TOP2A, which are involved in cell proliferation and epithelial-to-mesenchymal transition, are SUMOylated at several lysine residues by all PIAS ligases, suggesting a level of redundancy between these proteins. Confocal microscopy and biochemical experiments revealed that SUMOylation regulated TOP2A protein stability, while this modification is involved in the recruitment of Ki-67 nucleolar proteins containing the SUMO interacting motif. These results provide novel insights into both the redundant and specific regulatory mechanisms of cell proliferation and cell cycle mediated by PIAS SUMO E3 ligases.

Abstract IA015: Immunoprevention of triple negative breast cancer by TOP2A derived peptide vaccination

Abstract Top2A is a key enzyme involved in DNA replication and is a therapeutic target for several cancer types including breast cancer. Overexpression of Top2A has been observed in both human and mouse triple-negative breast cancer (TNBC). The present study evaluated both immunogenicity and antitumor efficacy of a newly formulated multi-peptide vaccine targeting multiple epitopes of the Top2A protein. Top2A-specific MHC II epitopes with optimal binding affinity were identified using a combined scoring system, which predicted their potential to elicit a Th1 immune response. The formulated vaccine contained top three Top2A peptides, which elicited the strongest immunologic response and showed 100% sequence homology between human and mouse. Anti-tumor efficacy of the Top2A vaccine was initially evaluated in a syngeneic TNBC mouse model, in which pre-graft preventive vaccination was associated with significantly decreased tumor growth as compared to the adjuvant controls. The Top2A peptide vaccine exhibited striking efficacy in a genetically engineered TNBC mouse model (C3(1)/Tag), reducing tumor burden by >90% when compared with adjuvant alone. Splenocytes collected from vaccinated animals showed a robust immunologic response to the immunizing peptides. There were no overt toxicities observed with the Top2A vaccination. To explore potential mechanisms underlying the anti-tumor response induced by Top2A vaccine treatment, scTCR-seq of tumors in both control and Top2A vaccine groups revealed new T cell clones as a consequence of Top2A vaccination. Furthermore, in vitro stimulation of these splenocytes by the vaccinated Top2A peptides resulted in the secretion of cytokines indicative of Th1 responses but with minimal secretion of Th2-related cytokines. Our data indicate that the newly developed multi-peptide Top2A vaccine is immunogenic and efficacious in the prevention of TNBC development and progression in vivo. Citation Format: Sang Beom Lee, Jing Pan, Donghai Xiong, Katie Palen, Bryon Johnson, Jeffrey E. Green, Shizuko Sei, Robert H. Shoemaker, Ronald A. Lubet, Yian Wang, Ming You. Immunoprevention of triple negative breast cancer by TOP2A derived peptide vaccination [abstract]. In: Proceedings of the Second Biennial NCI Meeting: Translational Advances in Cancer Prevention Agent Development (TACPAD); 2022 Sep 7-9. Philadelphia (PA): AACR; Can Prev Res 2022;15(12 Suppl_2): Abstract nr IA015.

ABCB1( P-glycoprotein) and TOP2A as predictive markers for anthracycline and taxane-containing chemotherapy regimens in neoadjuvant treatment of breast cancer

Topoisomerase II alpha (TOP2A) and protein has been shown to be a proliferation marker associated with tumor grade and Ki67 index. The predictive significance of TOP2A and ABCB1 together seems different among different subtypes of breast cancer. Our study evaluated the predictive impact of TOP2A and ABCB1 protein on breast cancer patients to whom prescribed neoadjuvant chemotherapy.

GINS1 promotes the proliferation and migration of glioma cells through USP15-mediated deubiquitination of TOP2A

SummaryGINS1 is a GINS complex subunit that functions along with the MCM2-7 complex and Cdc45 in eukaryotic DNA replication. Despite the significance of the GINS complex in the switch between quiescence and proliferation of glioma cells inside and outside the perinecrotic niche, the biological functions and the underlying mechanism of GINS1 remain unclear. Unlike in normal cells and tissues, GINS1 expression level was significantly upregulated in glioma cells and tissues. High expression of GINS1 predicted an advanced clinical grade and a poor survival. Functional assays revealed that GINS1 aggravated glioma malignant phenotypes in vitro and in vivo. Mechanistically, this study identified that GINS1 physically interacts with TOP2A. GINS1 promotes glioma cell proliferation and migration through USP15-mediated deubiquitination of TOP2A protein. Our results delineate the clinical significance of GINS1 in glioma and the regulatory mechanisms involved in glioma cell proliferation and migration. This work provides potential therapeutic targets for glioma treatment.

Oncogenetic Function and Prognostic Value of DNA Topoisomerase II Alpha in Human Malignances: A Pan-Cancer Analysis.

Increasing studies have revealed significant associations between TOP2A with oncogenesis and prognosis of human cancers; however, pan-cancer analysis has not been reported. Here, we explored the potential carcinogenic function and the association with clinical outcomes of TOP2A in 33 different human cancers. The results showed that TOP2A was amplified in 31 investigated cancers; TOP2A expression was significantly associated with metastasis of six different cancers and significantly associated with the survival of patients in ten different cancers; TOP2A-encoded protein was obviously upregulated in five available cancers; phosphorylated TOP2A protein at S1106 was significantly upregulated in all six available cancers. Moreover, TOP2A expression was found to be associated with the cancer-associated immune cell infiltration, including fibroblasts, Tregs, and macrophages. In addition, the Kyoto encyclopedia of genes and genomes (KEGG) pathway and Gene Ontology (GO) enrichment analyses revealed a most significant association between TOP2A with the Wnt signaling pathway and DNA conformation change. This work provides a comprehensive knowledge of TOP2A in different cancers, including carcinogenic function, prognostic values for metastasis, and clinical outcomes.

Expression analysis of p16 and TOP2A protein biomarkers in cervical cancer lesions and their correlation with clinico-histopathological characteristics in a referral hospital, Tanzania.

Biomarkers yield important information for early diagnosis of cervical cancer. However, they are rarely applied for prognosis of cervical cancer in Tanzania, where visual inspection assay with acetic acid or Lugol's iodine and Pap test are being used as the standard screening/ diagnostic methods. This was a retrospective hospital-based cross-sectional study that was conducted to assess cyclin-dependent kinase inhibitor (p16) and topoisomerase II-alpha (TOP2A) proteins expression among women seeking cervical cancer care at Kilimanjaro Christian Medical Centre, Tanzania between May 1, 2017 and May 10, 2018. Immunohistochemistry technique was used to detect the expressions of p16 and TOP2A proteins from the retrieved formalin-fixed and paraffin-embedded (FFPE) cervical biopsies. A total of 145 patients, with a mean age of 52.1 ± 12.9 years, were included in this study. Upon immunohistochemistry staining, 103 (71.0%) and 90 (62.1%) were p16 and TOP2A positive respectively. There was a strong association between histopathological class and p16/TOP2A expression levels (Fisher's exact test, p<0.001). Moreover, there was a strong positive correlation between p16/TOP2A and cancerous cervical lesions (Spearman's rank correlation coefficients = 0.833 and 0.687, p = 0.006 and 0.005, respectively). The age-adjusted odds ratio for predicting cervical cancer lesions were independently significant for p16/TOP2A biomarkers in FFPE cervical tissues [p16: OR = 1.142 (95% CI: 1.059-1.232, p<0.001) and TOP2A: OR = 1.046 (95% CI: 1.008-1.085, p = 0.015)]. Importantly, the diagnostic performance of p16 was higher than that of TOP2A in the diagnosis of cancerous lesions from non-cancerous cervical lesions (sensitivity: 97.2% versus 77.6%, accuracy: 92.8% versus 87.8%, respectively). Our study has highlighted that over-expression of TOP2A is related to the grade of cervical intraepithelial neoplasia but does not predict prognosis in cervical cancer. Similarly, expression of p16 is related to degree of histological dysplasia and malignancy, suggesting its prognostic and predictive value in the management of cervical cancers. Further bigger studies are needed to validate their applications in the early diagnosis of cervical cancer.

Abstract 1228: Novel small molecule inhibitors that target the exportin binding pocket of TOP2A for the treatment of multiple myeloma

Abstract Introduction Certain cancers have been shown to express a specific type of drug-resistance involving the export of drug target proteins from the nucleus of the cell to the cytoplasm. This mechanism of drug-resistance has been shown to be important in multiple myeloma (MM) and recently a nuclear export inhibitor, selinexor, has received approval by the FDA in MM. However, general nuclear export signal (NES) inhibitors like selinexor exhibit off-target toxicities when used in patients. We have begun to develop novel drugs that specifically prevent the export of the drug-target topoisomerase IIα (TOP2A). Methods In this study, we 1) generated an atomic homology model of human TOP2A to identify lead compounds targeting the NES of TOP2A; NES inhibitors (NESi), 2) examined binding specifically to TOP2A and inhibit nuclear export by immunofluorescence microscopy (IF) , nuclear-cytoplasmic fractionation, immunoprecipitation (IP), proximity ligation assay (PLA), and Biacore kinetic/affinity assay, and 3) tested the anticancer activity of these compounds in MM in vitro, ex vivo and in vivo MM. Results The atomic homology model of human TOP2A provided the basis for in silico molecular docking screenings of 139,735 small molecules (NCI) (MW &amp;lt; 500) to identify lead compounds targeting the NES of TOP2A. NESi compounds with the highest docking scores were assayed for their ability to induce apoptosis in human MM cells when used with the TOP2 inhibitor doxorubicin (DOX). We isolated four lead compounds that induced apoptosis when used with DOX and that inhibited nuclear export of TOP2A, as shown by IF and nuclear-cytoplasmic fractionation, IP, a PLA and Biacore kinetic/affinity assays. Inhibition by the lead compound, NCI-9138, was specific to TOP2A because p53 trafficking was unaffected and the inhibitor did not affect TOP2A protein expression or enzymatic function (decatenation). This NESi was found to sensitize human MM cells in vitro, in vivo mouse models, and patient MM cells exposed ex vivo to DOX but did not affect normal cell lines. These TOP2A-specific nuclear export inhibitors may potentially lead to a new approach in circumventing drug resistance. Conclusions Our previous studies have shown that TOP2A is exported from the nucleus of MM cells and that this is a significant cause of resistance to DOX therapy. In this study, we have shown that blocking export of TOP2A can sensitize human MM cells to DOX both in vitro, ex vivo in MM cells from patients, and in in vivo mouse models of MM. A drug that specifically inhibits the export of a single cancer-related protein has not been published. We intend to show that this unique approach to cancer-treatment may significantly advance the treatment of MM. This paper also demonstrates a proof of concept for the development of highly specific small molecule inhibitors that could be used as cancer therapeutics. Citation Format: Joel G. Turner, David Ostrov, Nicholas Lawrence, Jana Dawson, Juan Gomez, Alexis Bauer, Mark Meads, Alex Achille, Kenneth Shain. Novel small molecule inhibitors that target the exportin binding pocket of TOP2A for the treatment of multiple myeloma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1228.

MiRNA-145-5p expression and prospective molecular mechanisms in the metastasis of prostate cancer.

The clinicopathological implication and prospective molecular mechanisms of miRNA‐145‐5p in the metastasis of prostate cancer (PCa) stand unclear. Herein, it is found that miRNA‐145‐5p expression was remarkably reduced in 131 cases of metastatic PCa than 1371 cases of localised ones, as the standardised mean differences (SMD) was −1.26 and the area under the curve (AUC) was 0.86, based on miRNA‐chip and miRNA‐sequencing datasets. The potential targets of miRNA‐145‐5p in metastatic PCa (n = 414) was achieved from the intersection of miRNA‐145‐5p transfected metastatic PCa cell line data, differential expression of metastatic PCa upregulated genes and online prediction databases. TOP2A was screened as one of the target hub genes by PPI network analysis, which was adversely related to miRNA‐145‐5p expression in both metastatic PCa (r = −0.504) and primary PCa (r = −0.281). Gene‐chip and RNA‐sequencing datasets, as well as IHC performed on clinical PCa samples, showed consistent upregulated expression of TOP2A mRNA and protein in PCa compared with non‐PCa. The expression of TOP2A mRNA was also significantly higher in metastatic than localised PCa with the SMD being 1.72 and the AUC of sROC being 0.91. In summary, miRNA‐145‐5p may participate in PCa metastasis by binding TOP2A and be useful as a biomarker for the detection of metastatic PCa.

TOP2A expression predicts responsiveness to carfilzomib in myeloma and informs novel combinatorial strategies for enhanced proteasome inhibitor cell killing

Microarray was utilized to determine if a genetic signature associated with resistance to carfilzomib (CFZ) could be identified. Twelve human myeloma (MM) cell lines (HMCLs) were treated with CFZ and a cell-viability profile was assessed categorizing HMCLs as sensitive or resistant to CFZ. The gene expression profiles (GEP) of untreated resistant versus sensitive HMCLs revealed 29 differentially expressed genes. TOP2A, an enzyme involved in cell cycle and proliferation, was overexpressed in carfilzomib-resistant HMCLs. TOP2A protein expression levels, evaluated utilizing trephine biopsy specimens acquired prior to treatment with proteasome inhibitors, were higher in patients failing to achieve a response when compared to responding patients. Logistic-regression analysis confirmed that TOP2A protein expression was a highly significant predictor of response to PIs (AUC 0.738). Further, the combination of CFZ with TOP2A inhibitors, demonstrated synergistic cytotoxic effects in vitro, providing a rationale for combining topoisomerase inhibitors with CFZ to overcome resistance in MM.

Prognostic and predictive value of the combination of TOP2A and HER2 in node-negative tumors 2cm or smaller (T1N0) breast cancer.

We aim to evaluate the prognostic and predictive value of TOP2A and HER2 expression in T1N0 breast cancer patients. 299 cases with T1N0 breast cancer were obtained from the Oncomine database (Cohort 1) and 963 of T1N0 breast cancer patients from Sun Yat-sen Memorial Hospital (Cohort 2) were retrospectively enrolled. Kaplan-Meier product was applied to estimate survival curve. Cox proportional hazard models was used to identify prognostic factors. We used PSM (propensity score matching) to balance clinicopathologic characteristics among four groups of different HER2/TOP2A status. Survival between groups and chemotherapy regimens were analyzed, before and after PSM. In Cohort 1, we found that the group with HER2+ and higher expression of TOP2A mRNA was associated with poor breast cancer-specific survival (BCSS) compared to the group of HER2- with lower expression of TOP2A mRNA. In Cohort 2, HER2+ patients with higher TOP2A protein expression had greater risk of recurrence and distant recurrence compared to HER2- patients with lower expression of TOP2A protein. Among the patients who developed both HER2+ and higher expression of TOP2A protein and received chemotherapy, patients who received an anthracycline-based regimen had a significantly better recurrence-free survival (RFS) than those with a non-anthracycline-based regime. Patients with both HER2+ and high expression level of TOP2A protein predicts poor prognosis in T1N0 breast cancer patients. Patients with double positive for TOP2A protein and HER2 may benefit from anthracycline-based regimens.

A computational drug repositioning method applied to rare diseases: Adrenocortical carcinoma

Rare or orphan diseases affect only small populations, thereby limiting the economic incentive for the drug development process, often resulting in a lack of progress towards treatment. Drug repositioning is a promising approach in these cases, due to its low cost. In this approach, one attempts to identify new purposes for existing drugs that have already been developed and approved for use. By applying the process of drug repositioning to identify novel treatments for rare diseases, we can overcome the lack of economic incentives and make concrete progress towards new therapies. Adrenocortical Carcinoma (ACC) is a rare disease with no practical and definitive therapeutic approach. We apply Heter-LP, a new method of drug repositioning, to suggest novel therapeutic avenues for ACC. Our analysis identifies innovative putative drug-disease, drug-target, and disease-target relationships for ACC, which include Cosyntropin (drug) and DHCR7, IGF1R, MC1R, MAP3K3, TOP2A (protein targets). When results are analyzed using all available information, a number of novel predicted associations related to ACC appear to be valid according to current knowledge. We expect the predicted relations will be useful for drug repositioning in ACC since the resulting ranked lists of drugs and protein targets can be used to expedite the necessary clinical processes.

High expression of TOP2A gene predicted poor prognosis of hepatocellular carcinoma after radical hepatectomy.

BackgroundTopoisomerase (DNA) II alpha (TOP2A) is the up-regulated gene of the chromosome aggregation pathway. This gene encodes DNA topoisomerase, which can control and change the topological state of DNA during transcription and replication, participate in chromosome agglutination, separation, and relieve stress from kinking. Abnormally high expression of TOP2A is often associated with active cell proliferation. In studies of breast cancer, endometrial cancer, and adrenal cancer, it was found that high expression of TOP2A suggested a poor prognosis.MethodsA total of 15 pairs of fresh hepatocellular carcinoma (HCC) specimens and adjacent tissues were assembled, and the difference of TOP2A mRNA and protein expression level between tumor and adjacent tissues was detected by RT-PCR and Western blotting analysis, respectively. A total of 84 HCC paraffin specimens were selected for routine pathological grading. TOP2A expression was detected by immunohistochemistry (IHC). The correlation between TOP2A expression and clinicopathological features and survival time was analyzed.ResultsThe expression of TOP2A mRNA and protein in HCC tumor tissues was significantly higher than that in the adjacent tissues. IHC results indicated that the TOP2A positive rate in tumor tissues was significantly higher than that in adjacent tissues (84.52%, 71/84 vs. 8.33%, 7/84). High TOP2A expression was associated with poor tumor differentiation, significant cirrhosis, and larger tumor diameter. Among the patients whose survival time exceeded 60 m, 14 patients (37.84%, 14/37) had a high expression of TOP2A. Among the patients whose survival time was less than 60 m, the TOP2A expression was high in 37 cases (78.72%, 37/47). Kaplan-Meier survival analysis showed that the survival time of the group with low or no TOP2A expression was better than that of the group with high TOP2A expression (P=0.0011). Regression analysis of the clinical characteristics and 5-year survival follow-up data of 84 patients showed that cirrhosis and TOP2A high expression were the independent factors influencing the prognosis of HCC, while gender, age, tumor grade, hepatitis B virus (HBV) infection, and tumor size did not affect 5-year survival.ConclusionsThe high expression of TOP2A is associated with the invasiveness and poor prognosis of HCC tumors, and, together with liver cirrhosis, can be a prognostic indicator of radical HCC resection.

Abstract 3471: Mutual regulation of MDM4 and TOP2A in cancer cell proliferation

Abstract MDM4 and topoisomerase IIα (TOP2A) are important cancer-related factors. MDM4 acts as an oncoprotein promoting cancer progression by inhibiting tumor suppressor p53. As a DNA replication- and cell division-regulating enzyme, TOP2A is the main target of many anticancer therapy regimens, even though the exact role of TOP2A in cancer remains elusive. Herein we report that MDM4 and TOP2A bind to each other and are mutually upregulated at the post-translational level, leading to enhanced TOP2A catalytic activity, inhibition of p53, and increased tumor-cell proliferation. We demonstrate that the C-terminal region (CTR) of TOP2A binds to a unique sequence (residues 188-238) of MDM4, which contains an auto-inhibitory segment regulating MDM4-p53 interaction. TOP2A binding in turn activates MDM4 for p53 binding, resulting in enhanced inhibition of p53 and cancer cell proliferation. Conversely, binding of the MDM4 sequence to the CTR of TOP2A upregulates TOP2A protein expression. This in turn interferes with TOP2A DNA-decatenating activity. These results reveal novel functions of MDM4 and TOP2A as well as their interactions in oncogenesis, suggesting that inhibition of the MDM4-TOP2A interaction may represent a novel strategy in specifically and simultaneously targeting TOP2A and MDM4 for cancer treatment. Citation Format: Tao Liu, Hailong Zhang, Sha Yi, Lubing Gu, Muxiang Zhou. Mutual regulation of MDM4 and TOP2A in cancer cell proliferation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3471.

Mutual regulation of MDM4 and TOP2A in cancer cell proliferation.

MDM4 and topoisomerase IIα (TOP2A) are overexpressed in various human cancers. MDM4 acts as an oncoprotein which promotes cancer progression by inhibiting tumor suppressor p53. As a DNA replication‐ and cell division‐regulating enzyme, TOP2A is the main target of many anticancer therapy regimens; however, the exact role of TOP2A in cancer remains elusive. Herein, we report that MDM4 and TOP2A bind to each other and are mutually upregulated at the post‐translational level, leading to TOP2A protein stabilization, inhibition of p53, and increased tumor‐cell proliferation. We demonstrate that the C‐terminal region (CTR) of TOP2A binds to a unique sequence (residues: 188–238) of MDM4, which contains an auto‐inhibitory segment regulating the MDM4‐p53 interaction. TOP2A binding in turn activates MDM4 for p53 binding, resulting in enhanced inhibition of p53 and cancer cell proliferation. Conversely, binding of the MDM4 sequence to the CTR of TOP2A stabilizes TOP2A protein, leading to increased TOP2A protein expression. These results reveal novel functions of MDM4 and TOP2A as well as their interactions in oncogenesis, suggesting that inhibition of the MDM4‐TOP2A interaction may represent a novel strategy in specifically and simultaneously targeting TOP2A and MDM4 for cancer treatment.

Assessment of the concordance between fluorescence in-situ hybridization and immunohistochemistry in evaluating topoisomerase IIα in breast carcinoma

Background Topoisomerase IIα (TOP2A), encoded by TOP2A gene, is a molecular target for anthracycline therapy; hence, it can serve as a predictive or prognostic factor. There are several techniques used for measuring TOP2A status. The relative value of TOP2A gene amplification evaluated by fluorescence in-situ hybridization (FISH) and protein expression analyzed by immunohistochemistry (IHC) is yet unclear in the literature. Aim To evaluate the concordance between FISH and IHC for TOP2A status and to assess their relations with some prognostic clinicopathological parameters. Materials and methods A total of 86 invasive breast cancer paraffin blocks were retrieved from the archives of Pathology Department, National Cancer Institute, Cairo University, and were subjected to IHC, with a cutoff value of 10%, and FISH analysis with TOP2A/centromere enumeration probe 17 ratio of at least two for TOP2A expressions. The concordance between the results of both techniques as well as relations with selected prognostic parameters was evaluated. Results Of 86 invasive breast carcinomas, 43 (50%) revealed TOP2A protein overexpression, whereas 23 (26.7%) cases had TOP2A gene amplification. TOP2A gene amplification was recognized in 16 of 43 TOP2A protein-expressed cases, with 37.2% concordance rate and 0.209 κ statistics. TOP2A gene amplification was significantly correlated with human epidermal growth factor receptor-2 immuno-expressing tumors and with molecular subtypes (P Conclusion Discordance was evident between TOP2A gene amplification and protein expression. However, TOP2A gene amplification has a valuable prognostic influence, whereas protein expression is still a predictive factor useful to guide chemotherapy.

Topoisomerase II alpha and microtubule-associated protein-tau as a predictive marker in axillary lymph node positive breast cancer.

The aims of this study were to investigate the correlation between topoisomerase II alpha (TOP2A), microtubule-associated protein-tau (MAP-tau) and other prognostic factors in breast cancer and to evaluate the predictive value of TOP2A and MAP-tau in breast cancer patients who received anthracycline and taxane-containing adjuvant chemotherapy. Seventy patients with axillary lymph node positive breast cancer who underwent curative surgery between January 2000 and December 2005 were evaluated retrospectively. The levels of protein expression of TOP2A and MAP-tau were assessed using immunohistochemistry. Among the 70 patients, 43 (61.4%) showed TOP2A overexpression and 30 (42.9%) showed MAP-tau positivity. TOP2A overexpression was associated with p53 positivity and high histological grade. MAP-tau positivity was associated with a lower positive lymph node ratio, lower proliferative activity, and hormone receptor positivity. Based on the TOP2A and MAP-tau expression, there was no significant difference in disease-free survival in the breast cancer patients who received anthracycline and taxane-containing adjuvant chemotherapy. We conclude that immunohistochemical analysis of TOP2A and MAP-tau protein expression may not predict the benefits of adjuvant anthracycline and taxane chemotherapy in axillary node positive breast cancer.

The prognostic significance of topoisomerase II alpha protein in early stage luminal breast cancer

BackgroundTopoisomerase II alpha (TOP2A) protein has been shown to be a proliferation marker associated with tumor grade and Ki67 index. The prognostic effect of TOP2A seems different among different subtypes of breast cancer. The current study evaluated the prognostic impact of TOP2A protein on luminal breast cancer.MethodAltogether 434 stage I-II luminal breast cancer patients who underwent curative surgery in Sun Yat-Sen University Cancer Center between 2007 and 2009 were enrolled. TOP2A protein expression was assessed by immunohistochemistry. Clinical and pathological data were retrospectively collected.ResultWith a cut-off value of 30%, 127 (29.3%) patients were classified as TOP2A overexpression. TOP2A overexpression was associated with a higher tumor grade and Ki67 index. Patients with TOP2A high expression showed a significantly higher rate of distant metastasis and shorter distant metastasis free survival (DMFS) compared with patients with low TOP2A expression. The prognostic influence of TOP2A expression was more significant in years 5–8 after diagnosis, and more pronounced in stage II patients, luminal B disease, and patients treated with adjuvant endocrine therapy alone. Multivariate survival analysis revealed TOP2A overexpression was an independent fact for worse DMFS.ConclusionTOP2A protein showed a time dependent influence on prognosis in stage I-II luminal breast cancer, suggesting it might be a potential predictor of late recurrence for this group of patients.

Abstract P2-10-06: Identifying patients sensitive to anthracycline-containing therapy with quantitative proteomic and genomic profiling

Abstract Background: Selecting chemotherapy based on tumor biology can improve response rates and avert toxicity. Studies of the relationship between tumor expression of TOP2A protein and response to anthracycline-based chemotherapy have yielded contradictory results. Here we used mass spectrometry (MS) to evaluate associations between tumor molecular profiles and pathological complete response (pCR) in breast cancer patients treated with neoadjuvant anthracycline-containing therapy. Methods: Patients were selected from the ERNEST-B (Erlangen Neoadjuvant Study Breast), which is a retrospective cohort study. Archived tumor samples from anthracycline-treated breast cancer patients (n=133) were microdissected and solubilized. In each tumor sample, TOP2A and other target proteins were quantitated with a mass spectrometry assay. Molecular profiling also included RNA sequencing of the tumor and whole genome sequencing of both tumor and matched normal tissue sections. The cohort was dichotomized into high and low expressors of TOP2A using a protein level cutoff of 515 amol/ug of tumor protein. The difference in pCR (ypT0ypN0) rates between high and low expressors of TOP2A was assessed using a z-test for differences in proportion. Results: TOP2A protein was detected in 84 of 133 (63%) tumor samples from anthracycline-treated patients (range: 178 to 3044 amol/ug). Patients whose tumor expressed TOP2A protein above the cutoff of 515 amol/ug had higher pCR rates than patients with lower TOP2A expression (35.5% vs. 12.1%; odds ratio (OR): 4.48 [95% CI: 1.53-13.28], Fisher' exact test p = 0.004). The difference retained statistical significance in a logistic regression model adjusting for HR and HER2 status (OR: 2.23 [95% CI: 1.15-4.30], p=0.017). In a separate cohort of 19 breast cancer patients who did not receive anthracycline, there was no association between TOP2A protein expression level and pCR rate. Results from a validation cohort as well as the genomic analysis will be presented at the meeting. Conclusions: In a retrospective analysis of anthracycline-treated breast tumors, TOP2A expression was associated with a higher rate of pCR. Targeted proteomics may predict the response of breast cancer patients to anthracycline-based therapy. Citation Format: Schwartz S, Cecchi F, Tian Y, Yau C, Szeto C, Rübner M, Erber R, Beckmann MW, Hartmann A, Benz S, Hembrough T, Fasching P. Identifying patients sensitive to anthracycline-containing therapy with quantitative proteomic and genomic profiling [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr P2-10-06.

TOP2A and EZH2 Provide Early Detection of an Aggressive Prostate Cancer Subgroup.

Purpose: Current clinical parameters do not stratify indolent from aggressive prostate cancer. Aggressive prostate cancer, defined by the progression from localized disease to metastasis, is responsible for the majority of prostate cancer-associated mortality. Recent gene expression profiling has proven successful in predicting the outcome of prostate cancer patients; however, they have yet to provide targeted therapy approaches that could inhibit a patient's progression to metastatic disease.Experimental Design: We have interrogated a total of seven primary prostate cancer cohorts (n = 1,900), two metastatic castration-resistant prostate cancer datasets (n = 293), and one prospective cohort (n = 1,385) to assess the impact of TOP2A and EZH2 expression on prostate cancer cellular program and patient outcomes. We also performed IHC staining for TOP2A and EZH2 in a cohort of primary prostate cancer patients (n = 89) with known outcome. Finally, we explored the therapeutic potential of a combination therapy targeting both TOP2A and EZH2 using novel prostate cancer-derived murine cell lines.Results: We demonstrate by genome-wide analysis of independent primary and metastatic prostate cancer datasets that concurrent TOP2A and EZH2 mRNA and protein upregulation selected for a subgroup of primary and metastatic patients with more aggressive disease and notable overlap of genes involved in mitotic regulation. Importantly, TOP2A and EZH2 in prostate cancer cells act as key driving oncogenes, a fact highlighted by sensitivity to combination-targeted therapy.Conclusions: Overall, our data support further assessment of TOP2A and EZH2 as biomarkers for early identification of patients with increased metastatic potential that may benefit from adjuvant or neoadjuvant targeted therapy approaches. Clin Cancer Res; 23(22); 7072-83. ©2017 AACR.