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

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]

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.

BRCA1 and TOP2A gene amplification and protein expression in four molecular subtypes of breast cancer

We studied TOP2A amplification (using FISH methods), and TOP2A and BRCA1 protein overexpression (immunohistochemistry) in four molecular subtypes of breast cancer. Of 53 patients, 32 showed TOP2A and 38 showed BRCA1 overexpression. The highest percentage of TOP2A amplification (47.8%) and deletion (13%) was detected in Luminal B subtypes. Of 11 Luminal B tumors with TOP2A amplification, 9 (81.8%) overexpressed TOP2A. BRCA1 protein overexpression showed significant positive correlation with TOP2A protein expression. BRCA1 and TOP2A proteins exhibited similar patterns of expression in Luminal B and triple-negative breast cancer, suggesting the same prognosis in those patients.

HER2 amplification, HER2 overexpression,and TOP2A abnormalities in gastroesophageal cancers: Do they predict responsiveness to epirubicin-based chemotherapy?

e14672 Background: Abnormalities of TOP2A, a major target of anthracyclines, have been reported as positive predictive markers of response to anthracycline-based therapy in breast cancer. We determined abnormalities of TOP2A and HER2, and HER2 protein expression, in a cohort of patients with gastric or gastro-oesophageal junction adenocarcinomas treated with epirubicin-based chemotherapy. Methods: We studied 50 patients that received either adjuvant (74%) or perioperative (26%) epirubicin, cisplatin and 5-fluorouracil. All patients were screened for TOP2A amplification/deletion by fluorescent in situ hybridization (FISH), and for HER2 protein expression by immunohistochemistry (IHC). For each patient at least two paired surgical samples were examined. Tumors exhibiting a HER2 score of 2+ were further screened for gene amplification by FISH. We studied the association of TOP2A abnormalities and HER2 amplification/expression with recurrence free survival (RFS) and overall survival (OS). Results: By IHC, HER2 overexpression (score 3+) was detected in 9 patients. Intra-tumor heterogeneity of HER2 staining, resulting in discrepant HER2 scores, was detected in 4 of 50 (8%) paired samples analyzed. HER2 amplification was detected in 1 of 8 tumors with a HER2 score 2+. TOP2A amplification was observed in 1 of 50 tumors; none exhibited TOP2A deletions. In patients whose tumors showed HER2 overexpression or gene amplification (HER2-positive) and in patients with HER2-negative tumors, median RFS was 11.0 and 13.9 months, respectively (p= 0.35); median OS was 28.8 and 29.9 months, respectively (p= 0.56). A pathological complete response was observed in 1 of 4 HER2-positive patients that underwent perioperative chemotherapy. Conclusions: In this cohort of patients with early-stage gastroesophageal cancers who were treated with epirubicin-based chemotherapy, neither HER2 amplification nor HER2 overexpression were associated with outcome. TOP2A abnormalities are rare and their putative role as a determinant of responsiveness to anthracycline-based chemotherapy in gastroesophageal cancers is not supported by our findings.