TOP2A Amplification Research Articles

Neoadjuvant anthracycline-based (5-FEC) or anthracycline-free (docetaxel/carboplatin) chemotherapy plus trastuzumab and pertuzmab in HER2 + BC patients according to their TOP2A: a multicentre, open-label, non-randomized phase II trial

PurposePrevious studies have reported the benefit of dual HER2-targeting combined to neoadjuvant chemotherapy in HER2-amplified breast cancer (HER2 + BC). Moreover, besides the cardiac toxicity following their association to Trastuzumab, anthracyclines chemotherapy may not profit all patients. The NeoTOP study was designed to evaluate the complementary action of Trastuzumab and Pertuzumab, and the relevance of an anthracycline-based regimen according to TOP2A amplification status.MethodsOpen-label, multicentre, phase II study. Eligible patients were aged ≥ 18 with untreated, operable, histologically confirmed HER2 + BC. After centralized review of TOP2A status, TOP2A-amplified (TOP2A+) patients received FEC100 for 3 cycles then 3 cycles of Trastuzumab (8 mg/kg then 6 mg/kg), Pertuzumab (840 mg/kg then 420 mg/kg), and Docetaxel (75mg/m2 then 100mg/m2). TOP2A-not amplified (TOP2A-) patients received 6 cycles of Docetaxel (75mg/m2) and Carboplatin (target AUC 6 mg/ml/min) plus Trastuzumab and Pertuzumab. Primary endpoint was pathological Complete Response (pCR) using Chevallier’s classification. Secondary endpoints included pCR (Sataloff), Progression-Free Survival (PFS), Overall Survival (OS), and toxicity.ResultsOut of 74 patients, 41 and 33 were allocated to the TOP2A + and TOP2A- groups respectively. pCR rates (Chevallier) were 74.4% (95%CI: 58.9–85.4) vs. 71.9% (95%CI: 54.6–84.4) in the TOP2A + vs. TOP2A- groups. pCR rates (Sataloff), 5-year PFS and OS were 70.6% (95%CI: 53.8–83.2) vs. 61.5% (95%CI: 42.5–77.6), 82.4% (95%CI: 62.2–93.6) vs. 100% (95%CI: 74.1–100), and 90% (95%CI: 69.8–98.3) vs. 100% (95%CI: 74.1–100). Toxicity profile was consistent with previous reports.ConclusionOur results showed high pCR rates with Trastuzumab and Pertuzumab associated to chemotherapy. They were similar in TOP2A + and TOP2A- groups and the current role of neoadjuvant anthracycline-based chemotherapy remains questioned.Trial registration numberNCT02339532 (registered on 14/12/14).

Clinical Genetic Features and Neoadjuvant Chemotherapy Response in HER2-Low Breast Cancers: A Retrospective, Multicenter Cohort Study.

The objective of this work is to reveal differences in clinical and genetic features, as well as neoadjuvant chemotherapy (NAC) response, between HER2-low and HER2-zero or HER2-positive breast cancers. A total of 245 female patients with breast cancer were retrospectively enrolled from seven hospitals. Core needle biopsy (CNB) samples were collected before NAC and used for next-generation sequencing by a commercial gene panel. Clinical and genetic features, as well as NAC response, were compared between HER2-low and HER2-zero or HER2-positive breast cancers. The nonnegative matrix factorization (NMF) method was applied to cluster the C-Score of enrolled cases to reveal the intrinsic features of each HER2 subgroup. A total of 68 (27.8%) cases are HER2-positive, 117 (47.8%) cases are HER2-low, and 60 (24.5%) cases are HER2-zero. HER2-low breast cancers have a significantly lower pathologic complete response (pCR) rate than HER2-positive and HER2-zero breast cancers (p < 0.050 for all comparisons). Compared with HER2-low breast cancers, HER2-positive cases have higher rates of TP53 mutation, TOP2A amplification, and ERBB2 amplification, as well as lower rates of MAP2K4 mutation, ESR1 amplification, FGFR1 amplification, and MAPK pathway alteration (p < 0.050 for all comparisons). After clustering HER2-low cases by the NMF method, 56/117 (47.9%) are in cluster 1, 51/117 (43.6%) are in cluster 2, and 10/117 (8.5%) are in cluster 3. HER2-low cases in cluster 2 have the lowest pCR rate among the three clusters (p < 0.050). HER2-low breast cancers have significant genetic differences from HER2-positive cases. Genetic heterogeneity exists in HER2-low breast cancers and impacts on NAC response in this subgroup.

A DNA Repair and Cell Cycle Gene Expression Signature in Pediatric High-Grade Gliomas: Prognostic and Therapeutic Value.

Simple SummaryPediatric high-grade gliomas are incurable brain tumors for which there is a critical need for new therapeutic strategies as well as treatment-predictive biomarkers. This study examined the expression of DNA repair and cell cycle genes in pediatric high-grade gliomas with distinct driving mutations. The aim is to propose a novel classification of these tumors based on sub-groups exposing therapeutic vulnerabilities. Several DNA repair factors were identified that might become new diagnostic markers.Background: Pediatric high-grade gliomas (pHGGs) are the leading cause of mortality in pediatric neuro-oncology, displaying frequent resistance to standard therapies. Profiling DNA repair and cell cycle gene expression has recently been proposed as a strategy to classify adult glioblastomas. To improve our understanding of the DNA damage response pathways that operate in pHGGs and the vulnerabilities that these pathways might expose, we sought to identify and characterize a specific DNA repair and cell-cycle gene expression signature of pHGGs. Methods: Transcriptomic analyses were performed to identify a DNA repair and cell-cycle gene expression signature able to discriminate pHGGs (n = 6) from low-grade gliomas (n = 10). This signature was compared to related signatures already established. We used the pHGG signature to explore already transcriptomic datasets of DIPGs and sus-tentorial pHGGs. Finally, we examined the expression of key proteins of the pHGG signature in 21 pHGG diagnostic samples and nine paired relapses. Functional inhibition of one DNA repair factor was carried out in four patients who derived H3.3 K27M mutant cell lines. Results: We identified a 28-gene expression signature of DNA repair and cell cycle that clustered pHGGs cohorts, in particular sus-tentorial locations, in two groups. Differential protein expression levels of PARP1 and XRCC1 were associated to TP53 mutations and TOP2A amplification and linked significantly to the more radioresistant pHGGs displaying the worst outcome. Using patient-derived cell lines, we showed that the PARP-1/XRCC1 expression balance might be correlated with resistance to PARP1 inhibition. Conclusion: We provide evidence that PARP1 overexpression, associated to XRCC1 expression, TP53 mutations, and TOP2A amplification, is a new theranostic and potential therapeutic target.

Genomic correlates of response to adjuvant trastuzumab (H) and pertuzumab (P) in HER2+ breast cancer (BC): Biomarker analysis of the APHINITY trial.

1012 Background: We conducted a comprehensive genomic and immune-marker based analysis to identify prognostic and predictive biomarkers beyond clinical parameters (eg nodal status) in the setting of HP-based therapy. Methods: APHINITY, a phase III study, randomized 4805 pt with HER2+ BC to adjuvant chemo/H, plus P or placebo. Tumor Infiltrating Lymphocytes (TILs) were analysed using published methods. A nested case-control design with 1023 pt samples (3 controls matched to 1 distant relapse) underwent DNA (targeted) and RNA sequencing. Inverse probability weighting with Kaplan-Meier estimator weights, and models adjusted for treatment, hormone receptor, nodal status, age and chemotherapy type were used. Results are reported descriptively with 95% CIs. Interaction p-values are unadjusted for multiplicity. Results: DNA-seq, RNA-seq and TIL analyses were successful in 940/1023 (92%), 974/1023 (95%), and 4313/4804 (90%) samples, respectively. Prognosis (arms pooled): PI3K/PTEN/AKT alterations (HR 1.35; CI 1.01-1.79), MYC (HR 1.61; CI 1.16-2.23) or ZNF703 amplification (HR 1.62; CI 1.07-2.47) suggest poorer prognosis while TOP2A amplification (HR 0.49; CI, 0.32-0.74) suggests better prognosis independent of anthracycline use. Higher mRNA expression of an immune signature (IFNG, PD-L1, CXCL9 (HR 0.68; CI 0.52-0.89)) and TILs (HR 0.91; CI 0.86-0.96) also suggest better outcomes. Predictive effects: Table shows the HR for H+P benefit versus H for selected markers. PAM50 subtype did not predict benefit. Conclusions: In this comprehensive biomarker analysis, higher levels of immune markers and HER2 appeared to be associated with better prognosis and greater H+P benefit. Clinical trial information: NCT01358877. [Table: see text]

FISH Analysis of TOP2A and HER-2 Aberrations in Female Breast Carcinoma on Archived Material: Egyptian NCI Experience.

HER-2 and TOP2A genes are considered of great importance in breast cancer. Their copy number variability has been proposed to be a marker for the degree of chromosomal instability. Owing to the close proximity of TOP2A gene to HER-2 gene chromosome 17, TOP2A status is believed to affect therapeutic plan. The percentage of TOP2A aberrations is greatly variable among different studies. Is to investigate the relation between TOP2A and HER-2 gene amplification using fluorescence in situ hybridization technique. Archival blocks of 112 breast cancer Egyptian female patients were retrieved from the pathology department at NCI, Cairo University were retrieved and investigated using fluorescence in situ hybridization technique for TOP2A and HER-2 gene assessment. In addition, correlation with some clinicopathologic parameters was done. HER-2 gene amplification was encountered in about 33% of cases. TOP2A gene amplification and deletion were detected in 23.9% and 2.8% of studied cases. Moderate agreement was obtained between results of HER-2 gene and TOP2A gene amplification. HER-2 and TOP2A genes amplification are 2 separate genetic yet closely related events in breast cancer. Polysomy of chromosome 17 is proposed to be an early event in occurrence of TOP2A gene amplification. Further studies regarding effect of TOP2A gene in response to anthracyclines in Egyptian population should be planned for to establish its role in therapeutic planning.

Abstract P3-10-23: Changes in the expression of HER2 and other genes in HER2-positive metastatic breast cancer induced by treatment with ado-trastuzumab emtansine and/or pertuzumab/trastuzumab

Abstract Background: Although tremendous progress has been achieved with targeted therapy for HER2-positive (HER2+) metastatic breast cancer, most advanced tumors eventually develop resistance. Improving our understanding of mechanisms of resistance to anti-HER2 therapy is needed to develop new therapeutic approaches. The purpose of this study was to identify the mechanisms of resistance to treatment with ado-trastuzumab emtansine (T-DM1) and/or taxane/pertuzumab/trastuzumab (TPH). Methods: In our preclinical analysis, HER2+ cell lines resistant to treatment with T-DM1 (n=5), and pertuzumab/trastuzumab (n=3) were generated. HER2 expression in the original and resistant cell lines was compared using Western blot, and HER2 gene amplification was compared in them using fluorescence in situ hybridization (FISH) and a Droplet Digital Polymerase Chain Reaction HER2 copy-number-validation assay. In our clinical analysis, nine patients with HER2+ metastatic breast cancer who had progressed on T-DM1 and/or TPH were enrolled. Patients underwent biopsies following treatment with T-DM1 and/or TPH. Targeted next-generation sequencing was performed using the FoundationOne® assay (Foundation Medicine, Inc.) to identify gene alterations. Also, the HER2 expression before and after the therapy was compared using immunohistochemistry and/or FISH. Results: In preclinical analysis, HER2 expression/amplification by Western blot and gene copy-number analysis was significantly decreased in T-DM1–resistant cell lines (four of five cell lines; P &amp;lt; 0.01) but not in pertuzumab/trastuzumab-resistant cell lines (none of three cell lines). In clinical analysis, the patients' median age was 54 years (range, 45-77 years), and five patients (56%) were ER+. Five patients (56%) received first-line anti-HER2 therapy, and four patients (44%) received two lines of anti-HER2 therapy prior to enrollment. We observed loss of HER2 expression in four of nine patients (44%) after undergoing anti-HER2 therapy. After receiving TPH, one of eight patients (13%) lost HER2 positivity according to FISH. In contrast, after T-DM1, three of four tested patients (75%) lost HER2 amplification by FISH. As for next-generation sequencing, we analyzed seven samples: three after treatment with TPH and four after treatment with T-DM1. In four of these samples (57%), we observed loss of HER2 amplification: one after treatment with TPH and three after treatment with T-DM1. TP53 mutations were seen in all patients. Additionally, we observed TOP2A and MCL1 amplification in two patients with ERBB2 amplificationand AKT1 amplification in one patient with ERBB2 amplification loss. Conclusions: We show for the first time that T-DM1–resistant breast cancer cells lose HER2 expression and amplification. Additionally, we observed loss of HER2 expression in patient samples following treatment with HER2 targeted therapy. Further study of resistant tumor samples is required to understand the impact of HER2 loss on outcomes. For the time being, repeating biopsy analysis of a metastatic site after treatment with T-DM1 to determine the HER2 expression status is reasonable, and it may increase the efficacy of future anti-HER2 therapy. Citation Format: Kida K, Lee J, Liu H, Lim B, Murthy RK, Sahin AA, Tripathy D, Ueno NT. Changes in the expression of HER2 and other genes in HER2-positive metastatic breast cancer induced by treatment with ado-trastuzumab emtansine and/or pertuzumab/trastuzumab [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P3-10-23.

Abstract P3-04-09: Integrated analysis of genetic variations in Chinese breast cancer from a single institution

Abstract To investigate the characteristics of somatic mutation or copy number variations in Chinese breast cancer, tumor tissues from 105 patients diagnosed at age from 26 to 81 (median age 48) were assessed by next-generation sequencing technology using a customized panel, including 33 genes of putative tumor suppressors or oncogenes. At least one genetic alteration (including mutations, copy number variations and fusion genes) was observed in 99/105 (94%) samples. Similar to the previous report in TCGA dataset, TP53 (49%) and PIK3CA (43%) were the most frequently mutated genes, which occurred in a significant mutual exclusive manner (p&amp;lt;0.05). Three genes including MYC copy number amplification (35%), FGFR1 (19%) and GATA3 mutation (16%) were altered at the frequency of &amp;gt;10% in our dataset, in which the occurrence of MYC amplification was higher than the TCGA cohort (22%, p&amp;lt;0.05). Importantly, we identified four fusion genes of FGFR1 including one previously reported (TACC1-FGFR1) and three novel fusion ones (MIR1268A-FGFR1, LZTS1-AS1-FGFR1, and LINC01605-FGFR1). Unlike the high prevalence of CCND1 amplification (17%) and CDH1 mutations (13%) in TCGA dataset, genetic variations of CCND1 and CDH1 in our study occurred at a low frequency with 2% and 4%, respectively (p&amp;lt;0.05). In addition, we also identified three novel ESR1 mutations (ESR1 G74R, D230H and M250T) in the untreated patients with early breast cancer. Furthermore, nonnegative matrix factorization (NMF) clustering of genetic variations revealed five distinct molecular classes in our dataset. NMF class I was characterized by a high rate of HR+/ERBB2- tumors (80%) and genetic alterations of FGFR1 (100%). A gain of ERBB2 gene was observed in 93% of NMF class II along with TOP2A amplification in 57% of HR+/ERBB2+ tumors. NMF class III was characterized by a high rate of HR+/ERBB2- (95%) and GATA3 mutations (75%) without TP53 mutation. The characteristics of NMF class IV were the high rate of PIK3CA mutations (95%) and HR+/ERBB2- tumors (75%) along with low rate of TP53 mutations. More HR-/ERBB2- tumors (39%) were observed in NMF class V with a high rate of MYC amplification (82%) and TP53 mutation (89%). Further analysis in the TCGA cohort revealed the patients in NMF class V had the shorter survival time than other clusters. Collectively, we identified several novel genetic variations and generated a preliminary profile of somatic genetic aberrations that could classify Chinese breast cancer in this study, and may represent novel therapeutic targets for molecular subsets of breast cancer. This study was supported in part by National Natural Science Foundation of China (8160111571) and Guangdong Natural Science Foundation (2016A030313768). Citation Format: Liao N, Zhang G-C, Wang Y, Cao L, Li K, Ren C-Y, Wen L-Z, Shi Y, Zhu W, Chen X. Integrated analysis of genetic variations in Chinese breast cancer from a single institution [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 P3-04-09.

Molecular characterization of intestinal (IS) and diffuse subtypes (DS) of gastric adenocarcinomas.

60 Background: Gastric adenocarcinomas are histologically categorized into IS and DS. TCGA categorization suggests that IS is more common in tumors arising via the chromosomal instability pathway and DS is more common in genomically stable tumors. However, molecular differences between these subtypes are not well understood. Methods: Gastric adenocarcinomas were examined using NextGen sequencing (MiSeq platform on 47 genes or NextSeq on 592 genes), protein expression, and gene amplification techniques. For tumors sequenced with NextSeq, tumor mutational load (TML) was calculated based on somatic nonsynonymous missense mutations, and microsatellite instability (MSI) was evaluated on known MSI loci in target regions. Chi-square and t-tests were used for comparative analyses. Results: In total, 268 gastric adenocarcinomas with annotated histology (DS [n = 144]; IS [n = 124]) were analyzed. Patients with DS were younger than those with IS (mean age: 27y [DS] vs. 65y [IS], p &lt; 0.0001). The majority of patients with DS were female (56% [DS] vs. 35% [IS], p = 0.0004). Most frequently mutated genes in DS were TP53 (44%), ARID1A (37%) , CDH1 (14%), BAP1 (8%), and PIK3CA (5%); whereas the most frequent mutations in IS were ARID1A (57%), TP53 (51%), CDKN2A (13%), MUTYH (13%), and PIK3CA (12%). IS had a higher rate of APC mutations (10% vs. 2%, p = 0.04), whereas DS had a higher rate of CDH1 (14% vs. 2%, p = 0.01). There was no difference in PD-L1 tumor expression (DS: 4%; IS: 8%). IS, when compared to DS, exhibited higher overexpression of TOP2A (90% vs 50%, p &lt; 0.0001), TS (58% vs 28%, p &lt; 0.0001), RRM1 (47% vs 22%, p = 0.0006), and Her2/neu (14% vs 1%, p &lt; 0.0001), and greater Her2 amplification (19% vs 2%, p &lt; 0.0001). Microsatellite instability was not detected in DS, and there was a trend toward a higher frequency of MSI-high in IS (6% [IS] vs. 0% [DS], p = 0.06), as well as a higher mean TML (10.3 vs. 6.6 mutation/megabase, p = 0.01). Conclusions: Significant molecular differences between IS and DS gastric adenocarcinomas were observed, a finding that indicates different carcinogenic pathways and biology, as well as potential differences in response to therapy. Low frequency mutations in several druggable genes may provide therapeutic options.

Copy number variation and high expression of DNA topoisomerase II alpha predict worse prognosis of cancer: a meta-analysis.

Background: Increasing numbers of literatures have investigated the association between TOP2A and cancer prognosis. But the results of the relationship between the two were inconclusive. The aim of this meta-analysis was to elucidate whether TOP2A could predict prognosis of cancer.Materials and Methods: A systematically searching for potentially valuable literature was conducted through electronic databases containing PubMed and Web of Science. Hazard Ratio (HR) and their 95% confidence interval (CI) were used to assess the strength of association between TOP2A and cancer prognosis.Results: Finally twenty-five studies were included in this meta-analysis. High expression of TOP2A was associated with shorter disease free survival (DFS) of cancer prognosis compared with low expression of TOP2A (HR= 1.36, 95% CI= 1.18-1.57, P<0.001). Amplification of TOP2A gene showed no significant association with overall survival (OS), disease free survival (DFS) or relapse free survival (RFS) compared with non-amplification of TOP2A (OS: HR= 0.96, 95%CI= 0.75-1.22, P= 0.735; DFS: HR= 0.93, 95%CI= 0.70-1.23, P= 0.621; RFS: HR= 0.97, 95%CI= 0.71-1.34, P= 0.867). In the subgroup of regions, TOP2A amplification was associated with longer overall survival (HR= 0.66, 95%CI= 0.46-0.96, P= 0.029) in Australia. Alteration (amplification or deletion) of TOP2A gene demonstrated shorter survival according to OS and RFS compared with those with normal TOP2A status (OS: HR= 1.37, 95%CI= 1.22-1.55, P<0.001; RFS: HR= 1.26, 95%CI= 1.12-1.41, P<0.001).Conclusion: High TOP2A expression suggested significant relationship with worse cancer prognosis. Alteration (amplification or deletion) of TOP2A gene was also significantly related to shorter survival of cancer patients. Therefore, TOP2A might be used as an indicator for poor prognosis of cancer in the future.

HER2 and TOP2A Gene Amplification and Protein Expression in Upper Tract Urothelial Carcinomas.

HER2, a potential target for therapy, has been described to be amplified in urothelial carcinomas. As the topoisomerase II alpha (TOP2A) gene is located close to the HER2 gene on chromosome 17q12-q21, it is frequently either co-amplified or deleted with HER2 amplification. The purpose of this study was to assess the impact HER2 and TOP2A gene amplification as well as protein expression on outcomes of upper tract urothelial carcinoma (UTUC). HER2 and TOP2A gene amplification and protein expression were assessed in 81 patients with radical nephroureterectomy for UTUC. Immunohistochemistry and chromogenic in-situ hybridization was performed on formalin-fixed, paraffin-embedded samples. HER2 protein expression was observed in 27/81 (33%) cases, of which 8 cases exhibited amplification of HER2. One of them had an additional polysomy 17, whereas 6/67 HER2 non-amplified cases revealed a polysomy 17. Coamplification of HER2 and TOP2A was found in 4 cases, whereas 3 cases showed only HER2 amplification and 20 cases only TOP2A amplification. HER2 IHC overexpression was associated with higher-grade tumors (p=0.001), non-organ confined carcinomas (p=0.017), HER2 amplification (p<0.00001) and TOP2A amplification (p=0.016). HER2 amplification was association with higher tumor grade (p=0.001) and lymphnode metastasis (p=0.003). TOP2A IHC positivity was significantly associated with higher tumor grade (p=0.0004), TOP2A amplification (p=0.0003), polysomy 17 (p=0.035) and HER2 IHC overexpression (p=0.28), whereas all categories of tumor stage and HER2 amplification remained not related. TOP2A amplification was significantly more frequent in tumors with higher grade, higher tumor stage, polysomy 17 and distant metastasis (p=0.015; p=0.042; p=0.032; p = 0.011), respectively. In univariate analyses HER2 IHC positivity, TOP2A amplification, and polysomy 17 were associated with poor clinical outcome after surgery. HER2 IHC overexpression and TOP2A amplification are associated with features of biologically aggressive UTUC. Overexpression and/or amplification of HER2 and TOP2A could help identify patients who may benefit from targeted therapy.

Topoisomerase expression and amplification in solid tumours: Analysis of 24,262 patients

BackgroundTopoisomerase I (TOPO1) and topoisomerase IIα (TOP2A) are specific targets of multiple chemotherapy drugs. Increased expression of TOPO1 protein and amplification of the TOP2A gene have been associated with treatment response in colorectal and breast cancers, respectively. TOPO1 and TOP2A may be potential therapeutic targets in other malignancies as well. Summary of methodsWe analysed TOPO1 protein expression and TOP2A gene amplification in patients (n = 24,262 specimens) with diverse cancers. Since HER2 and TOP2A co-amplification have been investigated for predictive value regarding anthracycline benefit, we analysed specimens for HER2 amplification as well. ResultsOverexpressed TOPO1 protein was present in 51% of the tumours. Four percent of the tumours had TOP2A amplification, with gallbladder tumours and gastroesophageal/oesophageal tumours having rates over 10%. Overall, 4903 specimens were assessed for both TOP2A and HER2 amplification; 129 (2.6%) had co-amplification. High rates (>40%) of HER2 amplification were seen in patients with TOP2A amplification in breast, ovarian, gastroesophageal/oesophageal and pancreatic cancer. ConclusionOur data indicate that increased TOPO1 expression and TOP2A amplification, as well as HER2 co-alterations, are present in multiple malignancies. The implications of these observations regarding sensitivity to chemotherapy not traditionally administered to these tumour types merits investigation.

Amplification of HER2 and TOP2A and deletion of TOP2A genes in a series of Taiwanese breast cancer.

The prognostic relevance of topoisomerase II alpha (TOP2A) copy number change remains not well established. This study is aimed to investigate the frequency and pattern of TOP2A aberrations; to correlate TOP2A alterations with human epidermal growth factor receptor 2 (HER2) status and clinicopathological parameters, and further to explore prognostic value of TOP2A and HER2 status in breast cancer in Taiwan. We analyzed tissue samples from 311 invasive carcinomas in tissue microarrays for TOP2A and HER2 status by fluorescent in situ hybridization. TOP2A copy number change is an infrequent genetic event (9.8% amplification and 2.7% deletion) and is present in both HER2-amplified and nonamplified tumors. TOP2A amplification is statistically associated with age >50 at diagnosis (P = 0.016) and HER2 amplification (P < 0.001). HER2 amplification, but not TOP2A amplification, is a predictor of unfavorable prognosis (P = 0.002). Univariate and multivariate analysis showed that higher histologic grading, positive nodal involvement, and HER2 positivity were associated with poorer overall survival. Cytogenetically, double minutes-type amplification is the predominant pattern for both genes (HER2: 64% and TOP2A: 93.1%). Homogeneous staining region-type signals of both genes are resistant to RNase digestion, supporting that these were not nuclear accumulation of mRNA transcripts. Our results demonstrate the prognostic value of tumor grading, nodal involvement, and HER2 status in Taiwanese breast cancer. TOP2A aberrations are an infrequent event independent of HER2 status, and TOP2A amplification carries no prognostic value. The predictive value of TOP2A aberrations in patients of breast cancer taking athracycline-containing treatment in Taiwan remains to be determined in prospectively well-designed clinical trials.

Abnormality of TOP2A expression and its gene copy number variations in neuroblastic tumors

Objective: To detect TOP2A protein expression and gene copy number alterations, and to analyze related clinical and pathological implications in pediatric neuroblastic tumors (NT). Methods: Immunohistochemistry was used to detect TOP2A protein expression. Fluorescence in situ hybridization (FISH) was used to detect numerical aberrations of TOP2A. Results: TOP2A protein was expressed in 59.1%(52/88) of cases, which was associated with differentiation (P=0.006), Ki-67 index (P<0.01) and MKI (P=0.001). Twenty-eight cases (35.0%, 28/88) showed TOP2A gene amplification, which was correlated with the age (P<0.01), clinical stage (P=0.028), high risk group (P=0.001), Ki-67 index (P=0.040) and differentiation (P=0.014). Survival analysis showed that TOP2A expression was related to survival rate. Multivariate analyses showed that TOP2A expression was an independent predictor for poor prognosis (P=0.010). Conclusions: More than half of the cases show TOP2A expression, which is more likely associated with NB, high Ki-67 index and high MKI. Cases with TOP2A expression have shorter survivals and poorer prognosis. TOP2A amplification is seen in 35% and likely occurs in patients older than 18 months and at advanced INSS stages (Ⅲ and Ⅳ). As a target of the anthracycline-based adjuvant drugs, TOP2A test can be used to select patient with NT for the therapy.

TOP2A copy number and TOP2A expression in uterine benign smooth muscle tumours and leiomyosarcoma

AimsTo examine TOP2A copy number, TOP2A expression, and its prognostic value in uterine leiomyosarcoma (LMS) and other benign smooth muscle tumours.MethodsWe analysed 37 patients treated for uterine LMS with immunohistochemistry...

Predictive Biomarker Profiling of > 6000 Breast Cancer Patients Shows Heterogeneity in TNBC, With Treatment Implications

BackgroundTriple-negative breast cancer (TNBC) is an aggressive disease without established targeted treatment options for patients with metastatic disease. This study was undertaken to evaluate potentially actionable biomarkers in a large cohort of TNBC and compare them with non-TNBCs. Materials and MethodsWe evaluated 6341 (2111 TNBC and 4230 non-TNBC) breast cancer samples at a central laboratory for biomarkers of potential drug response across multiple platforms, including gene sequencing, protein expression, and gene copy number. ResultsTNBC expresses androgen receptor (AR) in a significantly (P < .05) lower percentage of cases (17%) than hormone receptor (HR)-positive and human epidermal growth factor receptor 2 (HER2)-positive breast carcinomas (59% and 79%, respectively), and gene comutations were differentially associated with AR-positive versus AR-negative cases. Higher AR expression levels in TNBC predicted for lower Ki-67 levels. Seventy percent of TNBC harbored a phosphatidylinositol-4,5-bisphosphate 3-kinase, catalytic subunit alpha (PIK3CA), v-akt murine thymoma viral oncogene homolog 1 (AKT1), or phosophatase and tensin homolog (PTEN) aberration. TNBC patients had a significantly lower PIK3CA mutation rate (13%) than all other subtypes (P < .05) and a higher tumor protein p53 (TP53) mutation rate (64%) than the estrogen receptor (ER)-positive cases (approximately 30%; P < .05). Topoisomerase 2 (TOP2A) amplification was observed in 1.3% of TNBC and in 1.6% of HER2-negative, HR-positive cancers; in contrast, HER2-positive, HR-negative or HR-positive cancers exhibited TOP2A amplification in 19% and 40% of cases, respectively (P <.05). ConclusionMulti-platform molecular profiling identifies subgroups of TNBC with different biomarker profiles, suggesting numerous potentially targetable alterations in TNBC. TNBC is further characterized by different gene mutations and proliferative activity relative to AR expression, highlighting a need for comprehensive pathologic examination with potential to develop different, individualized treatment options.

TOP2A amplification and overexpression in hepatocellular carcinoma tissues.

Hepatocellular carcinoma (HCC) is the leading cause of cancer death in men worldwide owing to limited insights into pathogenesis and unsatisfactory efficacy of current therapies. HER2 and TOP2A genes are coamplified in breast and some other cancers. In this study, we investigated gene aberrations of HER2 and TOP2A and protein expressions of HER2, TOP2A, Ki-67, and p53 in tumor and matched nontumor tissues, as well as their associations with clinicopathological features. Gene aberrations were evaluated by FISH and protein expressions by IHC. Neither HER2 overexpression nor HER2 gene amplification was observed in both tumor tissues and matched nontumor tissues. By contrast, TOP2A overexpression was detected in 72.5% of tumor tissues but not detected in matched nontumor tissues. However, TOP2A gene amplification was not observed in both tumor and matched nontumor tissues. TOP2A overexpression was significantly associated with HCC tumor tissues (P < 0.001), hepatitis B surface antigen (HBsAg) in the serum (P = 0.004), and Ki-67 (P = 0.038) but not with age, tumor size, alpha-fetoprotein, TP53, and copy number of TOP2A gene and chromosome 17 centromere. In conclusion, TOP2A overexpression in HCC was not secondary to gene amplification. In addition, neither HER2 amplification nor overexpression could be used as prognostic and predictive marker in HCC.

Chromosome 17 Centromere Duplication and Responsiveness to Anthracycline-Based Neoadjuvant Chemotherapy in Breast Cancer

Human epidermal growth factor receptor 2 (HER2) and topoisomerase II alpha (TOP2A) genes have been proposed as predictive biomarkers of sensitivity to anthracycline chemotherapy. Recently, chromosome 17 centromere enumeration probe (CEP17) duplication has also been associated with increased responsiveness to anthracyclines. However, reports are conflicting and none of these tumor markers can yet be considered a clinically reliable predictor of response to anthracyclines. We studied the association of TOP2A gene alterations, HER2 gene amplification, and CEP17 duplication with response to anthracycline-based neoadjuvant chemotherapy in 140 patients with operable or locally advanced breast cancer. HER2 was tested by fluorescence in situ hybridization and TOP2A and CEP17 by chromogenic in situ hybridization. Thirteen patients (9.3%) achieved pathologic complete response (pCR). HER2 amplification was present in 24 (17.5%) of the tumors. TOP2A amplification occurred in seven tumors (5.1%). CEP17 duplication was detected in 13 patients (9.5%). CEP17 duplication correlated with a higher rate of pCR [odds ratio (OR) 6.55, 95% confidence interval (95% CI) 1.25-34.29, P = .026], and analysis of TOP2A amplification showed a trend bordering on statistical significance (OR 6.97, 95% CI 0.96-50.12, P = .054). TOP2A amplification and CEP17 duplication combined were strongly associated with pCR (OR 6.71, 95% CI 1.66-27.01, P = .007). HER2 amplification did not correlate with pCR. Our results suggest that CEP17 duplication predicts pCR to primary anthracycline-based chemotherapy. CEP17 duplication, TOP2A amplifications, and HER2 amplifications were not associated with prognosis.

HER2 and TOP2A amplification in a hospital-based cohort of breast cancer patients: associations with patient and tumor characteristics

Gene amplification is an important factor for altered gene expression in breast cancers. TOP2A-amplification often occurs concomitantly with HER2 amplification, and it has been suggested to be predictive for the response to anthracycline chemotherapy. This study assessed the correlation between HER2 status and TOP2A co-amplification, the possible association of TOP2A single-nucleotide polymorphisms with the frequency of this co-amplification as well as confirmation of association with outcome. HER2 and TOP2A amplification were analyzed in a tissue microarray from a clinical cohort study. Additionally, a common genetic variant (rs13695) in the TOP2A gene was genotyped in germline DNA. HER2 gene amplification was compared with HER2-IHC findings assessed during clinical routine work, and the association between all the biomarkers analyzed and the clinical outcome was determined. As an exploratory aim, rs13695 genotypes were compared with TOP2A amplification status. HER2 amplification was seen in 101 of 628 (16.1 %) and TOP2A amplification in 32 (5.1 %) cancers. No TOP2A amplification occurred without HER2 co-amplification. HER2 amplification was found in 8, 13.6, and 55.1 % of patients with HER2-IHC 0/1+, 2+, and 3+ tumors, respectively. HER2-IHC was not associated with an effect on the prognosis, but HER2-FISH was. There was an association between the rs13695 genotype and TOP2A amplification status (P = 0.03). Although there was a significant correlation between HER2 status determined by IHC and HER2 by FISH, only HER2 gene amplification status by FISH was correlated with outcome indicating greater utility for FISH in routine clinical settings.

Prognostic and predictive value of copy number alterations in invasive breast cancer as determined by multiplex ligation-dependent probe amplification

Breast cancer is a leading cause of morbidity and mortality in women worldwide. About 70% of breast cancers are estrogen receptor (ER) positive. Blocking estrogen action by tamoxifen has been the treatment of choice in ER positive breast cancers for more than 30years. In the past, several studies have revealed associations between gene copy number alterations and responsiveness to tamoxifen therapy, but so far no single gene copy number alteration could completely explain the response variation observed between individual breast cancer patients. Here, we set out to perform a simultaneous analysis of copy number alterations of several genes involved in the prognosis and response to therapy by multiplex ligation-dependent probe amplification (MLPA). A case-control study was designed encompassing 170 non-metastatic ER positive breast cancer patients (case group = 85, control group = 85). All patients in the control group had received standard adjuvant tamoxifen treatment for 5years without any evidence of recurrence. Patients in the case group had experienced early recurrences while receiving tamoxifen treatment. 76% of the patients of the case group and 73% of the patients of the control group had received anthracycline-based adjuvant chemotherapy. Gene copy number alterations detected by MLPA in both groups were compared. Amplification of CCND1 (OR = 3.13; 95% CI = 1.35 to 7.26; p = 0.006) and TOP2A (OR = 3.05; 95% CI = 1.13 to 8.24; p = 0.022) were significantly more prevalent in the case group, compared to the control group. In a multivariate analysis CCND1 (p = 0.01) and TOP2A (p = 0.041) amplifications remained significant predictors of recurrence. Our results indicate that CCND1 amplification may serve as a useful biomarker for hormone responsiveness, and that TOP2A amplification may serve as a useful prognostic biomarker.

Abstract PD4-1: Comparison of mutations and protein expression in potentially actionable targets in 5500 triple negative vs. non-triple negative breast cancers

Abstract Introduction Triple negative breast cancer is a heterogeneous disease with no established targeted treatment options for patients with metastatic disease. Methods A cohort of 5500 patient samples was evaluated for differences and similarities in gene mutation (Sanger or Illumina Truseq), protein expression (immunohistochemistry), and/or amplification/rearrangement (CISH or FISH) between triple negative and non-triple negative breast cancers, with 16% of those patients identified as triple negative by IHC testing. Results PIK3CA and P53 gene mutations, PIK3CA, TOP2A, and EGFR amplification (measured as ≥ 4 copies in 40% or more tumor cells by FISH), and AR, EGFR, SPARC, and TS protein expression were shown to differ between the triple negative and non-triple negative breast cancer patients. The rate of mutation of the P53 tumor suppressor gene, measured by sequencing was twice as high in TNBC patients as in non-TNBC patients (60% versus 30%), while PIK3CA was mutated in half as many TNBC patients (12% versus 31% of non-TNBC patients). Amplification of Her2, PIK3CA, cMYC, and TOP2A was significantly greater in the non-TNBC patient population, while amplification of EGFR was significantly higher in the TNBC patient population (24% versus 13% of non-TNBC patients). Androgen receptor was expressed in 56% of the non-TNBC cohort but in only 15% of the TNBC cohort. Conclusions The gene mutation and protein expression differences in the triple negative subset of breast cancer patients, such as EGFR amplification, SPARC protein expression and P53 gene mutation, could inform new therapeutic possibilities for these difficult to treat, often more aggressive cancers. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr PD4-1.