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Article

A DNA Damage Response Gene Panel for Different Histologic Types of Epithelial Ovarian Carcinomas and Their Outcomes

by
Ying-Cheng Chiang
1,2,
Po-Han Lin
3,4,
Tzu-Pin Lu
5,
Kuan-Ting Kuo
6,
Yi-Jou Tai
2,7,
Heng-Cheng Hsu
7,8,
Chia-Ying Wu
2,7,
Chia-Yi Lee
7,8,
Hung Shen
7,8,
Chi-An Chen
1,2,*,† and
Wen-Fang Cheng
1,2,7,9,*,†
1
Department of Obstetrics and Gynecology, College of Medicine, National Taiwan University, Taipei 100226, Taiwan
2
Department of Obstetrics and Gynecology, National Taiwan University Hospital, Taipei 100226, Taiwan
3
Department of Medical Genetics, National Taiwan University Hospital, Taipei 100226, Taiwan
4
Graduate Institute of Medical Genomics and Proteomics, College of Medicine, National Taiwan University, Taipei 100025, Taiwan
5
Institute of Epidemiology and Preventive Medicine, Department of Public Health, National Taiwan University, Taipei 100025, Taiwan
6
Department of Pathology, College of Medicine, National Taiwan University, Taipei 100225, Taiwan
7
Graduate Institute of Clinical Medicine, College of Medicine, National Taiwan University, Taipei 100225, Taiwan
8
Department of Obstetrics and Gynecology, National Taiwan University Hospital, Hsin-Chu Branch, Hsin-Chu 30059, Taiwan
9
Graduate Institute of Oncology, College of Medicine, National Taiwan University, Taipei 100025, Taiwan
*
Authors to whom correspondence should be addressed.
†
Equal contribution.
Biomedicines 2021, 9(10), 1384; https://doi.org/10.3390/biomedicines9101384
Submission received: 10 August 2021 / Revised: 20 September 2021 / Accepted: 29 September 2021 / Published: 3 October 2021
(This article belongs to the Special Issue Mechanisms and Novel Therapeutic Approaches for Gynecologic Cancer)
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Abstract

:
DNA damage response (DDR) is important for maintaining genomic integrity of the cell. Aberrant DDR pathways lead to accumulation of DNA damage, genomic instability and malignant transformations. Gene mutations have been proven to be associated with epithelial ovarian cancer, and the majority of the literature has focused on BRCA. In this study, we investigated the somatic mutation of DNA damage response genes in epithelial ovarian cancer patients using a multiple-gene panel with next-generation sequencing. In all, 69 serous, 39 endometrioid and 64 clear cell carcinoma patients were enrolled. Serous carcinoma patients (69.6%) had higher percentages of DDR gene mutations compared with patients with endometrioid (33.3%) and clear cell carcinoma (26.6%) (p < 0.001, chi-squared test). The percentages of DDR gene mutations in patients with recurrence (53.9 vs. 32.9% p = 0.006, chi-squared test) or cancer-related death (59.2 vs. 34.4% p = 0.001, chi-squared test) were higher than those without recurrence or living patients. In endometrioid carcinoma, patients with ≥2 DDR gene mutations had shorter PFS (p = 0.0035, log-rank test) and OS (p = 0.015, log-rank test) than those with one mutation or none. In clear cell carcinoma, patients with ≥2 DDR gene mutations had significantly shorter PFS (p = 0.0056, log-rank test) and OS (p = 0.0046, log-rank test) than those with 1 DDR mutation or none. In the EOC patients, somatic DDR gene mutations were associated with advanced-stage tumor recurrence and tumor-related death. Type I EOC patients with DDR mutations had an unfavorable prognosis, especially for clear cell carcinoma.

1. Introduction

Epithelial ovarian carcinoma (EOC) is a major cause of death in women worldwide, and patients are usually diagnosed at an advanced stage with a 5-year survival of less than 50% [1,2,3,4]. Clinical prognostic factors include cancer stage, histological subtypes, tumor grade, residual tumor size after debulking surgery and response to chemotherapy. Despite an initial good response to primary treatments of debulking surgery and adjuvant platinum-based chemotherapy, the majority of patients experience a cancer relapse that is resistant to salvage treatments and eventually die of the disease [4,5].
Precision medicine is the current direction for cancer management depending on the specific genetic or molecular features of cancer. There are several subtypes of EOC—high-grade serous, clear cell, endometrioid, mucinous and low-grade serous—that could be viewed as distinct diseases for their differences in clinical course and pathological features [6,7]. To date, the most promising target therapies for EOC are anti-angiogenesis agents and poly ADP-ribose polymerase inhibitors (PARPi). Bevacizumab in combination with chemotherapy has demonstrated improved progression-free survival, and an overall survival benefit in high-risk patients [8,9,10]. Maintenance therapy with PARPi has revised the management of EOC in newly diagnosed and recurrent diseases. The identification of BRCA mutations or homologous recombination deficiency (HRD) status is critical for selecting potential patients, but both positive and negative patients as defined by current HRD assays benefited from PARPi [11,12,13,14,15].
DNA damage response (DDR) is important for maintaining a cell’s genomic integrity, and the DDR pathway is composed of various molecules that detect DNA damage, activate cell-cycle checkpoints, trigger apoptosis, and coordinate DNA repair [16,17,18]. Several exogenous or endogenous sources (e.g., oxidative damage, radiation, ultraviolet light, cytotoxic materials, replication errors) may result in DNA damage that may eventually lead to genomic instability and cell death [19]. DDR consists of several pathways, including base excision (BER), mismatch (MMR) and nucleotide excision repair (NER); translesion synthesis (TLS) for single-strand break repair; homologous recombination (HR) and nonhomologous DNA end joining (NHEJ) for double-strand break repair; and cell cycle regulation (CCR) (27, 28). Homologous recombination is an error-proof repair pathway to restore the original sequence at the double-strand DNA break. BRCA 1/2 genes participating in HR and maintaining PARPi therapy for BRCA-mutated EOC is a good example of synthetic lethality [20]. Several other DDR genes have been identified as potential targets for novel cancer therapy under clinical investigation [16,17]. Understanding the complex DDR pathways is helpful for exploring the feasibility of novel DDR inhibitors in clinical practice. In the study, we investigated the somatic mutations of DDR genes in 172 EOC patients using a targeted DDR gene panel using a next-generation sequencing method. The correlation of the somatic DDR gene mutations, clinical parameters and outcomes was analysed.

2. Materials and Methods

2.1. Patients and Specimens

The study protocol was approved by the National Taiwan University Hospital Research Ethics Committee (201509042RINA, approved on 24 November 2015 and 201608025RINA, approved on 07 October 2016). Informed consent from all participants was obtained and the methods were performed in accordance with the guidelines and regulations. From December 2015 to October 2018, 172 women diagnosed with epithelial ovarian cancer who had received debulking surgery and adjuvant chemotherapy were enrolled. The cancerous tissue specimens collected during debulking surgery were immediately frozen in liquid nitrogen and stored at −70 °C. A portion of the tissue specimens were sent for pathological examinations to confirm the diagnosis and ensure tumorous tissue sufficient for the following experiments. Clinical data were obtained from medical records, including age, cancer stage, the findings during debulking surgery, treatment course and recurrence. Optimal debulking surgery was defined as a maximal residual tumor size <1 cm following surgery. The tumor grade based on International Union Against Cancer criteria, and cancer stage was based on International Federation of Gynecology and Obstetrics (FIGO) criteria [21]. All patients received platinum-based adjuvant chemotherapy and regular follow-ups after primary treatments. Recurrence was defined as abnormal results from imaging studies (including computerized tomography or magnetic resonance imaging), elevated CA-125 (more than twice the upper normal limit) for two consecutive tests in 2-week intervals, or a biopsy-proven disease. Progression-free survival (PFS) was defined as the time from the date of primary treatment completion to the date of confirmed recurrence, disease progression or last follow-up. Overall survival (OS) was defined as the period from surgery to the date of death related to EOC or the date of last follow-up.

2.2. The Panel of DNA Damage Repair Genes

We selected 60 genes involved in DNA damage response (DDR) for the gene panel (Table 1), including genes of homologous recombination (HR), nonhomologous DNA end joining (NHEJ), base excision repair (BER), mismatch repair (MMR), nucleotide excision repair (NER), translesion synthesis (TLS) and cell cycle regulation (CCR) [16,17].

2.3. Genomic DNA Extraction

Genomic DNA was isolated using a QIAGEN Genomic DNA extraction kit according to the manufacturer’s instructions (Qiagen Inc., Valencia, CA, US). The purity and concentration of the genomic DNA were checked by agarose gel electrophoresis and the OD260/280 ratio.

2.4. Library Preparation, Next-Generation Sequencing, and Sequence Mapping

The genomic DNA was fragmented with Covaris fragmentation protocol (Covaris, Inc., Woburn, MA, US). The size of the fragmented genomic DNA was checked by Agilent Bioanalyzer 2100 (Agilent Technologies, Inc., Santa Clara, CA, US) and NanoDrop spectrophotometer (Thermo Fisher Scientific, Inc., Wilmington, DE, US). The target gene library was generated with NimblGen capture kits (Roche NimblGen, Inc. Hacienda Dr Pleasanton, CA, US). The samples were sequenced by Illumina MiSeq with paired-end reads of 300 nucleotides.
The analysis algorithm was conducted according to our previous protocol [22]. Briefly, the raw sequencing data were aligned with the reference human genome (Feb. 2009, GRCh37/hg19) with Burrows–Wheeler Aligner software (version 0.5.9) [23]. SAM tools (version 0.1.18) was used for data conversion, sorting, and indexing [24]. For single nucleotide polymorphisms (SNPs) and small insertion/deletions (indels), Genome Analysis Toolkit (GATK; version 2.7) was used for variant calling with Base/indel-calibrator and HaplotypeCaller. Pindel or Breakdancer software were used for structural variants larger than 100 bp which cannot be identified by GATK, such as large deletions, insertions and duplications [25]. After variant calling, ANNOVAR was used for annotation of the genetic variants [26,27]. The dbSNP, Exome sequencing Project 6500 (ESP6500) and the 1000 Genomes variant dataset were used to filter common variants of sequencing results.

2.5. Variant Classification

The sequence variants were classified according to the IARC variant classification [28]. The pathogenic mutations were defined as large-scale deletion, frame-shift mutation, nonsense mutation, genetic variants associated with uncorrected splicing and mutations affecting protein function demonstrated by functional analyses. The pathogenic and likely pathogenic mutations were used as deleterious mutations in our study. An allele frequency greater than 0.01 in the general population in the 1000 Genomes variant dataset or ESP6500 database were considered benign or likely benign genetic variants. Silent and intronic variants that did not affect splicing were also considered benign or likely benign. Other variants, mainly missense mutations without known functional data, were considered as variants of uncertain significance (VUSs). To reduce their number, bioinformatics analyses, including PolyPhen2 and SIFT, were used to evaluate potential pathogenicity [29,30,31]. The VUSs were suspected of being deleterious mutations if they met two criteria: (1) a population frequency of less than 0.01 in the 1000 Genomes and ESP6500 databases and (2) a bioinformatics analysis result with a SIFT score less than 0.05 and a polyphen2 score greater than 0.95.

2.6. Statistical Analysis

All statistical analyses were performed using the Statistical Package for Social Sciences software package (IBM SPSS Statistics for Windows, Version 22.0. IBM Corp. Armonk, NY, US) and R (version 3.1.2, The R Foundation for Statistical Computing, Institute for Statistics and Mathematics, Wirtschaftsuniversität Wien, Welthandelsplatz Vienna, Austria). One-way ANOVA was used to compare continuous variables and a chi-squared test was used for categorical variables. Survival curves were generated using the Kaplan–Meier method, and differences were calculated using the log-rank test. A multivariate Cox’s regression model was used to evaluate the prognostic factors for progression-free survival (PFS) and overall survival (OS). Statistical significance was set as a p value of less than 0.05.

3. Results

3.1. Clinical Characteristics of the Patients

There were 172 EOC patients enrolled: 69 serous, 39 endometrioid and 64 clear cell carcinomas (Table 2). There were 68 high-grade serous carcinomas (type II tumor) and 104 type I tumors. The median age was 52, and the median pre-treatment CA125 value was 400 U/mL; 59.9% were diagnosed at an advanced cancer stage, and 65.1% had undergone optimal debulking surgery; 59.3% had disease recurrence, and 44.2% died of EOC. All patients received adjuvant platinum and paclitaxel chemotherapy.

3.2. Deleterious DDR Gene Mutations

As shown in Table 3, 114 deleterious somatic mutations were identified from 26 genes of our 60-gene DDR panel in 78 EOC patients: 27 nonsense mutations in 23 patients, 28 frameshift mutations in 20, 28 missense mutations in 26 patients and 31 mutations involving uncorrected splicing in 29 patients. There were single-gene mutations in 57 patients, and multiple-gene mutations in 21: 2 mutations in 14 patients, 3 mutations in 2, 4 mutations in 3, 5 mutations in 1 and 6 mutations in 1 patient (Figure 1). We also identified 109 missense mutations classified as variants of uncertain significance (VUSs) with the potential of being deleterious mutations after searching the database (http://www.ncbi.nlm.nih.gov/snp, accessed on 28 September 2021) and bioinformatic analyses (Table S1 and Figure S1).
The pattern of prevalent mutated DDR genes was different among the histological subtypes (Figure 1). The proportion of wild type DDR genes was 54.7% in all EOC patients; 30.4% in serous carcinoma, 66.7% in endometrioid carcinoma and 73.4% in clear cell carcinoma. The top three prevalent mutated DDR genes were TP53 (27.9%), MUTYH (6.4%) and BRCA2 (5.8%) for all patients. Serous carcinoma—TP53 (56.5%), BRCA2 (5.8%) and RAD51C (5.8%); endometrioid carcinoma—TP53 (15.4%), ATM (12.8%) and MSH2 (7.7%); clear cell carcinoma—MUTYH (9.4%), TP53 (4.7%), BRCA2 (3.1%) and ERCC8 (3.1%). The top three prevalent mutated subgroups of DDR genes were CCR (30.8%), HR (10.5%) and BER (7.0%) for all patients. Serous carcinoma—CCR (58.05%), HR (15.9%) and BER (5.8%); endometrioid carcinoma—CCR (23.1%), MMR (15.4%) and HR (7.7%); clear cell carcinoma—BER (9.4%), CCR (6.3%) and HR (6.3%). For detailed information, please refer to Table S2 and Figure S2.

3.3. Correlation of DDR Gene Mutations with Clinical Outcomes of the EOC Patients

We evaluated the correlations between the mutation of DDR genes, the clinicopathologic parameters and outcome of the EOC patients. As shown in Table 4, type II tumors had a higher percentage of HR gene mutations than type I tumors (16.18 vs. 6.73%, p = 0.048, chi-squared test). Endometrioid carcinoma (15.38%) had a higher percentage of MMR mutations than those of serous carcinoma (2.90%) and clear cell carcinoma (4.69%) (p = 0.03, chi-squared test). Low-grade tumors had a higher percentage of MMR mutations compared with high-grade tumors (17.24 vs. 4.20%, p = 0.009, chi-squared test). Type II tumors had a higher percentage of DSBR mutations than type I tumors (17.65 vs. 6.73%, p = 0.026, chi-squared test). Serous carcinoma (57.97%) had a higher percentage of CCR mutations than those of endometrioid carcinoma (23.08%) and clear cell carcinoma (6.25%) (p < 0.001, chi-squared test). Type II tumors had higher percentage of CCR mutations than those of type I tumors (58.82 vs. 12.50%, p < 0.001, chi-squared test). The advanced-stage patients had a higher percentage of CCR mutations than the early-stage patients (42.72 vs. 13.04%, p < 0.001, chi-squared test). The recurrent patients had a higher percentage of CCR mutations than those without recurrence (39.22% vs. 18.57%, p = 0.004, chi-squared test). Patients who died of EOC had higher percentages of CCR mutations than living patients (40.79 vs. 22.92%, p = 0.012, chi-squared test). Serous carcinoma (69.57%) had higher percentage of DDR mutations than those of endometrioid carcinoma (33.33%) and clear cell carcinoma (26.56%) (p < 0.001, chi-squared test). Type II tumors had a higher percentage of DDR mutations than type I tumors (70.59 vs. 28.85%, p < 0.001, chi-squared test). The advanced stage patients had higher percentage of DDR mutations than the early-stage patients (57.28 vs. 27.54%, p < 0.001, chi-squared test). Recurring patients had a higher percentage of DDR mutations than those without recurrence (53.92 vs. 32.86%, p = 0.006, chi-squared test). Patients who died of EOC had a higher percentage of DDR mutations than living patients (59.21 vs. 34.38%, p = 0.001, chi-squared test).
EOC patients without DDR gene mutation had longer progression-free survival (PFS) (p = 0.0072, log-rank test, Figure 2A) and overall survival (OS) (p = 0.022, log-rank test, Figure 2B) than those with 1 DDR or ≥2 DDR mutations. In serous carcinoma, patients with or without DDR mutations had similar PFS (p = 0.56, log-rank test, Figure 2C). Patients with ≥2 DDR mutations had a trend of better OS than those with 1 mutation or none, but it was not statistically significant (p = 0.47, log-rank test, Figure 2D). In endometrioid carcinoma, patients with ≥2 DDR gene mutations had shorter PFS (p = 0.0035, log-rank test, Figure 2E) and OS (p = 0.015, log-rank test, Figure 2F) than those with 1 mutation or none. In clear cell carcinoma, patients with ≥2 DDR gene mutations had significantly shorter PFS (p = 0.0056, log-rank test, Figure 2G) and OS (p = 0.0046, log-rank test, Figure 2H) than those with 1 DDR mutation or none.
Tumor recurrence with CCR gene mutation (HR: 1.68 (1.12–2.50), p = 0.011), 1 DDR gene mutation (HR: 1.71 (1.12–2.60), p = 0.013), endometrioid carcinoma (HR: 0.17 (0.08–0.37), p < 0.001), type II tumor (HR: 2.69 (1.81–4.00), p < 0.001), advanced-stage carcinoma (HR: 5.29 (3.16–8.85), p < 0.001), high-grade tumor (HR: 5.57 (2.26–13.70), p < 0.001) and optimal debulking surgery (HR: 0.28 (0.18–0.41), p < 0.001) were significant in the univariate Cox regression model (Table 5). Advanced-stage carcinoma (HR: 3.08 (1.63–5.80), p = 0.001) and optimal debulking surgery (HR: 0.51 (0.32–0.80), p = 0.004) were important prognostic factors in the multivariate analysis. Cancer-related death with TLS gene mutation (HR: 33.76 (3.95–289.00), p = 0.001), 1 DDR gene mutation (HR: 1.96 (1.20–3.20), p = 0.007), endometrioid carcinoma (HR: 0.12 (0.04–0.38), p < 0.001), type II tumor (HR: 1.88 (1.19–2.96), p = 0.007), advanced-stage carcinoma (HR: 6.84 (3.28–14.25), p < 0.001), high-grade tumor (HR: 17.97 (2.50–129.29), p = 0.004) and optimal debulking surgery (HR: 0.26 (0.16–0.41), p < 0.001) were significant in the univariate Cox regression model. Type II tumor (HR: 0.35 (0.20–0.60), p < 0.001), TLS gene mutation (HR: 9.57 (1.08–84.83), p = 0.042), advanced-stage carcinoma (HR: 4.82 (2.09–11.09), p < 0.001) and optimal debulking surgery (HR: 0.38 (0.22–0.64), p < 0.001) were important prognostic factors in the multivariate analysis.

4. Discussion

Our study showed that nearly half of the epithelial ovarian cancer (EOC) patients had DNA damage response (DDR) gene mutations with varied proportions of histological subtypes. Two-thirds of serous adenocarcinoma patients, one-third of endometrioid adenocarcinoma patients and one-fourth of clear cell carcinoma patients had DDR gene mutations. Our DDR gene panel consisted of the genes involved in single-strand break repair, double-strand break repair and cell cycle regulation, including the genes recommended by National Comprehensive Cancer Network (NCCN) guidelines as cost-effective tools for assessing the lifetime risk of EOC, such as ATM, BRCA1/2, BRIP1, MLH1, MSH2, MSH6, PALB2, RAD51C and RAD51D [32]. The major components of DDR gene mutations were CCR in serous, CCR and SSBR in endometrioid and SSBR in clear cell carcinomas; CCR and DSBR in type II tumors (high-grade serous carcinoma in the cohort); and SSBR in type I tumors. A multiple DDR gene panel increased the detection rate of somatic mutation of genes involved in DNA damage repair pathway in comparison with a BRCA test alone. The percentage of BRCA 1/2 somatic mutation in serous carcinoma was 7.2, which was compatible with the 6–7% in previous studies [33,34,35,36,37]. The non-BRCA HR somatic mutation of our study was more than 10% in serous and endometrioid carcinomas, and the MMR somatic mutation was around 15% in endometrioid carcinomas, which was compatible with the previous study [38].
Our study showed that ovarian clear cell carcinoma patients with DDR gene mutations had an unfavorable survival prognosis. Those who had somatic DDR mutations were significantly associated with advanced-stage carcinomas, tumor recurrence and tumor-related death. The trend was different in histological subtypes as serous carcinomas or type II tumors with DDR mutation showed a better survival trend. Non-serous or type I EOC patients with DDR mutations had a poor prognosis, especially in clear cell carcinoma. Ovarian clear cell carcinoma is an aggressive drug-resistant subtype of EOC in association with endometriosis and glycogen accumulation. It accounts for about 5–13% of all EOCs in Western populations, but up to 20–25% in East Asia, including Taiwan [2,3]. Previous studies showed that the somatic mutations of ovarian clear cell carcinoma (mainly in ARID1A, PIK3CA, KRAS and PPP2R1A) might be related to chromatin remodeling, cell proliferation, cell cycle checkpointing and cytoskeletal organization [39,40,41,42,43,44,45,46,47,48,49]. However, the frequent mutations of ARID1A, PIK3CA, PPP2R1A or TP53 in ovarian clear cell carcinoma did not correlate well with the prognosis [45]. Other infrequent gene mutations of clear cell carcinoma included ARID1B, ARID3A, CREBBP, CSMD3, CTNNB1, LPHN3, LRP1B, MAGEE1, MLH1, MLL3, MUC4, PIK3R1, PTEN and TP53 [41,43,46,48,49]. DDR gene mutations in ovarian clear cell carcinoma was unclear in the literature, and our finding of an unfavorable prognosis in clear cell carcinoma patients with DDR gene mutations could provide useful information.
Our DDR gene panel could provide a scientific rationale for patient selection in future clinical trials that target DNA damage repair response pathways, especially in clear cell carcinoma. BRCA gene tests or companion HRD assays are currently suggested for PARPi, but there are unmet problems that need to be resolved [11,12,13,14,15,20]. The most important one is that the HRD assays cannot consistently identify patients who do not benefit from PARPi therapy. The consensus for the cut-off value was indeterminate because the thresholds of HRD assays were developed from retrospective exploratory analyses [11,50,51]. Generally, advance-stage, high-grade serous carcinoma patients with tumor BRCA (tBRCA) mutations, including germline (gBRCA) or somatic (sBRCA), derived the greatest benefit from PARPi maintenance therapy [11,12,13,14,15]. Approximately 11–18% of patients had a gBRCA mutation, and another 6–7% patients had an sBRCA mutation with a negative gBRCA test [33,34,35,36,37]. However, about 5% of gBRCA mutated patients tested negative for tBRCA [52,53,54]. The non-BRCA HR gene mutations were usually pooled together to interpret the association with clinical outcomes in previous studies because of their relatively low prevalence [35,55,56,57]. Twenty-one platinum-sensitive recurrent patients with non-BRCA somatic mutations (BRIP1, CDK12, RAD54L and RAD51B) derived benefit from olaparib in study 19 [58]. In ARIEL2, there were 20 patients with non-BRCA HR gene mutations (ATM, BRIP1, CHEK2, FANCA, FANCI, FANCM, NBN, RAD51B, RAD51C and RAD54L), but the sensitivity in discriminating a rucaparib response was only 11% [59]. However, BRCA wild type EOC patients still benefitted from PARPi, which indicated that a BRCA test by itself was inadequate for selecting EOC patients for PARPi [13,14,15]. It needs to be determined which individual or panel of non-BRCA HR genes could be used to predict a PARPi response, especially in non-serous EOC patients.
There were limitations to our study. First, germline gene mutations were not investigated. These not only inform the patients but also identify family members of the possible risk of malignancy [52,53,54]. The NCCN suggested germline gene tests of ATM, BRCA1/2, BRIP1, MLH1, MSH2, MSH6, PALB2, RAD51C, RAD51D and STK11 to assess the lifetime risk of EOC [32], but how many genes should be included in the panel is inconclusive. Second, the numerous variants of uncertain significance (VUSs) identified by multiple gene panels would cause controversy in risk assessment and management [60,61,62]. The biological functions and clinical impacts of most individual mutations in the genomic loci have not been well characterized, especially for VUSs [63]. Even in the well-studied BRCA gene, there is a difference among laboratories in the VUS reporting rate (3–50%), detection protocols and management strategies [64]. Further sharing and integration of gene sequencing data in an open database might decrease VUSs. Third, the cohort sample was not large enough; only the trends of clinical prognosis that correlated with each DDR pathway were found. Further large-scale investigations are needed.

5. Conclusions

Our study found that nearly half of the EOC patients had DDR gene mutations of varying proportions in the histological subtypes. Patients with somatic DDR mutations were significantly associated with advanced-stage carcinoma, tumor recurrence and tumor-related death. Type I EOC patients with DDR mutations had an unfavorable prognosis, especially for clear cell carcinoma. A broad multiple-gene DDR panel would provide not only comprehensive information of gene mutations but also a rationale for a future study of a novel therapy target for DNA damage response pathways.

Supplementary Materials

The following are available online at https://www.mdpi.com/article/10.3390/biomedicines9101384/s1, Figure S1: Variants of uncertain significance (VUS) with the potential of being deleterious mutations, Figure S2: The distribution of all 114 deleterious DDR gene mutations, Table S1: Variants of uncertain significance (VUSs) with the potential of being deleterious mutations in the cohort, Table S2: The percentages of DDR gene mutations in all patients.

Author Contributions

Conceptualization, Y.-C.C., P.-H.L., W.-F.C. and C.-A.C.; methodology, Y.-C.C. and P.-H.L.; software, P.-H.L. and T.-P.L.; validation, Y.-C.C., P.-H.L. and T.-P.L.; formal analysis, Y.-C.C., P.-H.L. and T.-P.L.; investigation, Y.-C.C., Y.-J.T., H.-C.H., C.-Y.W., C.-Y.L., H.S.; resources, Y.-C.C., K.-T.K., Y.-J.T., H.-C.H., C.-Y.W., C.-Y.L., H.S., C.-A.C. and W.-F.C.; data curation, Y.-C.C., P.-H.L. and T.-P.L.; writing—original draft preparation, Y.-C.C. and P.-H.L.; writing—review and editing, C.-A.C. and W.-F.C.; visualization, Y.-C.C. and P.-H.L.; supervision, C.-A.C. and W.-F.C.; funding acquisition, Y.-C.C., C.-A.C. and W.-F.C. All authors have read and agreed to the published version of the manuscript.

Funding

This work was supported by research grants from the National Taiwan University Hospital (NTUH. 105-N02, UN105-059, 108-S4230 and 109-S4570).

Institutional Review Board Statement

The study was conducted according to the guidelines of the Declaration of Helsinki, and approved by the Ethics Committee of National Taiwan University Hospital (201509042RINA, approved on 24 November 2015 and 201608025RINA, approved on 07 October 2016).

Informed Consent Statement

Informed consent was obtained from all subjects involved in the study.

Data Availability Statement

The datasets generated and/or analyzed during the current study are available from the corresponding author on reasonable request.

Acknowledgments

The authors thank the A1 laboratory of the National Taiwan University Hospital for the Illumina Miseq NGS platform, the National Applied Research Laboratories for providing access of high-performance computer to analyze NGS data, and the 7th Core Laboratory Facility of the Department of Medical Research of National Taiwan University Hospital for supporting the work.

Conflicts of Interest

The authors declare no conflict of interest.

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Figure 1. Deleterious DNA damage response (DDR) gene mutations in 172 epithelial ovarian carcinoma (EOC) patients (A) The pattern of DDR mutations of different histologic subtypes. (B) The percentages of DDR mutations in all 172 EOC patients. (C) The percentages of DDR mutations in different histologic subtypes. (D) The percentages of DDR mutations classified by different pathways in different histologic subtypes.
Figure 1. Deleterious DNA damage response (DDR) gene mutations in 172 epithelial ovarian carcinoma (EOC) patients (A) The pattern of DDR mutations of different histologic subtypes. (B) The percentages of DDR mutations in all 172 EOC patients. (C) The percentages of DDR mutations in different histologic subtypes. (D) The percentages of DDR mutations classified by different pathways in different histologic subtypes.
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Figure 2. Kaplan–Meier analysis of progression-free survival (PFS) and overall survival (OS) in 172 epithelial ovarian carcinoma (EOC) patients. (A) PFS of 172 EOC patients. Note: EOC patients with DDR gene mutation(s) had shorter PFS (p = 0.0072, log-rank test). (B) OS of 172 EOC patients. Note: EOC patients with DDR gene mutation(s) had shorter OS (p = 0.022, log-rank test) (C) PFS of 69 serous carcinoma patients. Note: Serous carcinoma patients with ≥2 DDR gene mutations had a trend of better PFS although no statistical significance. (D) OS of 69 serous carcinoma patients. Note: Serous carcinoma patients with ≥2 DDR gene mutations had a trend of better OS although no statistical significance. (E) PFS of 39 endometrioid carcinoma patients. Note: Endometrioid carcinoma patients with ≥2 DDR gene mutations had poor PFS (p = 0.0035, log-rank test). (F) OS of 39 endometrioid carcinoma patients. Note: Endometrioid carcinoma patients with ≥2 DDR gene mutations had poor OS (p = 0.014, log-rank test). (G) PFS of 64 clear cell carcinoma patients. Note: Clear cell carcinoma patients with ≥2 DDR gene mutations had significantly shorter PFS (p = 0.0056, log-rank test). (H) OS of 64 clear cell carcinoma patients. Note: Clear cell carcinoma patients with ≥2 DDR gene mutations had significantly shorter OS (p = 0.0046, log-rank test).
Figure 2. Kaplan–Meier analysis of progression-free survival (PFS) and overall survival (OS) in 172 epithelial ovarian carcinoma (EOC) patients. (A) PFS of 172 EOC patients. Note: EOC patients with DDR gene mutation(s) had shorter PFS (p = 0.0072, log-rank test). (B) OS of 172 EOC patients. Note: EOC patients with DDR gene mutation(s) had shorter OS (p = 0.022, log-rank test) (C) PFS of 69 serous carcinoma patients. Note: Serous carcinoma patients with ≥2 DDR gene mutations had a trend of better PFS although no statistical significance. (D) OS of 69 serous carcinoma patients. Note: Serous carcinoma patients with ≥2 DDR gene mutations had a trend of better OS although no statistical significance. (E) PFS of 39 endometrioid carcinoma patients. Note: Endometrioid carcinoma patients with ≥2 DDR gene mutations had poor PFS (p = 0.0035, log-rank test). (F) OS of 39 endometrioid carcinoma patients. Note: Endometrioid carcinoma patients with ≥2 DDR gene mutations had poor OS (p = 0.014, log-rank test). (G) PFS of 64 clear cell carcinoma patients. Note: Clear cell carcinoma patients with ≥2 DDR gene mutations had significantly shorter PFS (p = 0.0056, log-rank test). (H) OS of 64 clear cell carcinoma patients. Note: Clear cell carcinoma patients with ≥2 DDR gene mutations had significantly shorter OS (p = 0.0046, log-rank test).
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Table
Table 1. List of the DNA damage response (DDR) gene panel.
Table 1. List of the DNA damage response (DDR) gene panel.
GeneDDR PathwayGeneDDR Pathway
ATMCCRku70/XRCC6NHEJ
BARD1HRku80/XRCC5NHEJ
BRCA1HRMDM4CCR
BRCA2/FANCD1HRMLH1MMR
BRIP1/FANCJHRMLH3MMR
CHEK2CCRMRE11HR
DDB1NERMSH2MMR
DDB2NERMSH3MMR
ERCC1NERMSH6MMR
ERCC2/XPDNERMUTYHBER
ERCC3/XPBNERNBNHR
ERCC4NERNBS1HR
ERCC5/BIVMNEROGG1BER
ERCC6/CSBNERPMS1MMR
ERCC8/CSANERPMS2MMR
FANCAHRPOLD1TLS
FANCBHRPOLETLS
FANCCHRPOLBTLS
FANCD1/BRCA2HRPOLHTLS
FANCD2HRPOLKTLS
FANCEHRRAD50HR
FANCFHRRAD51HR
FANCG/XRCCHRRAD51C/FANCOHR
FANCIHRRAD51DHR
FANCJ/BRIP1HRTP53CCR
FANCL/PHF9HRXPANER
FANCMHRXPCNER
FANCN/PALB2HRXRCC2NHEJ
FANCO/RAD51CHRXRCC3NHEJ
FANCP/SLX4HRXRCC4NHEJ
Note: BER: base excision repair; CCR: cell cycle regulation; DDR: DNA damage repair; HR: homologous recombination; MMR: mismatch repair; NER: nucleotide excision repair; NHEJ: nonhomologous DNA end joining; TLS: translesion synthesis.
Table
Table 2. Characteristics of the epithelial ovarian cancer patients.
Table 2. Characteristics of the epithelial ovarian cancer patients.
Patient Numbers172
Median Age (years old)52 (29–85)
Median CA 125 (U/mL)400 (12–7265)
Histology
Serous carcinoma69 (40.1%)
Endometrioid carcinoma39 (22.7%)
Clear cell carcinoma64 (37.2%)
FIGO stage
Early69 (40.1%)
Advanced103 (59.9%)
Grade
Low29 (16.9%)
High143 (83.1%)
Debulking surgery
Optimal112 (65.1%)
Suboptimal60 (34.9%)
Recurrence
Yes102 (59.3%)
No70 (40.7%)
Death
Yes76 (44.2%)
No96 (55.8%)
Table
Table 3. The deleterious DDR gene mutations in the patients.
Table 3. The deleterious DDR gene mutations in the patients.
GeneMutationTranscriptgDNA/cDNAAmino AcidReported/Novel
ATMframeshift deletionNM_000051c.1402_1403delp.K468fsrs587781347
ATMframeshift deletionNM_000051c.8426delAp.Q2809fsrs587782558
ATMframeshift insertionNM_000051c.4736dupAp.Q1579fsrs864622164
ATMmissense mutationNM_000051c.C6200Ap.A2067Drs397514577
ATMnonsense mutationNM_000051c.C5188Tp.R1730Xrs764389018
ATMnonsense mutationNM_000051c.C850Tp.Q284Xrs757782702
BARD1frameshift insertionNM_000465c.70_71insGTp.P24fsNA
BRCA1nonsense mutationNM_007294c.3531dupTp.S1178_K1179delinsXNA
BRCA1nonsense mutationNM_007294c.G2635Tp.E879Xrs80357251
BRCA2frameshift deletionNM_000059c.1585delTp.F529fsNA
BRCA2frameshift insertionNM_000059c.7407dupTp.T2469fsrs397507916
BRCA2nonsense mutationNM_000059c.4965delCp.Y1655Xrs80359475
BRCA2nonsense mutationNM_000059c.A5623Tp.K1875XNA
BRCA2nonsense mutationNM_000059c.C2590Tp.Q864Xrs1060502414
BRCA2nonsense mutationNM_000059c.C6952Tp.R2318Xrs80358920
BRCA2nonsense mutationNM_000059c.G3922Tp.E1308Xrs80358638
BRIP1frameshift insertionNM_032043c.394dupAp.T132fsrs587781416
CHEK2splicingNM_007194g. 29130716 C>GNA
ERCC8frameshift deletionNM_000082c.191_195delp.S64fsNA
ERCC8splicingNM_000082c.1123-2->TNA
ERCC8splicingNM_000082c.1123-2->TNA
ERCC8splicingNM_000082c.1123-2->TNA
ERCC8splicingNM_000082c.1123-2->Trs777444521
FANCCnonsense mutationNM_000136c.G1225Tp.E409XNA
FANCGsplicingNM_004629c.511-2->CNA
FANCIsplicingNM_001113378c.3187-2A>GNA
FANCMframeshift deletionNM_020937c.3998delAp.Q1333fsrs746983128
MLH1frameshift deletionNM_000249c.1771delGp.D591fsNA
MLH1splicingNM_000249c.2104-2A>Grs267607889
MLH1splicingNM_000249c.790+2T>Crs267607790
MLH3missense mutationNM_001040108c.G2221Tp.V741Frs28756990
MLH3missense mutationNM_001040108c.G2221Tp.V741Frs28756990
MLH3missense mutationNM_001040108c.G2221Tp.V741Frs28756990
MLH3missense mutationNM_001040108c.G2221Tp.V741Frs28756990
MRE11frameshift insertionNM_005590c.1222dupAp.T408fsrs774440500
MSH2nonsense mutationNM_000251c.C226Tp.Q76Xrs63750042
MSH2nonsense mutationNM_000251c.G1738Tp.E580Xrs63751411
MSH2splicingNM_000251c.943-1G>Crs12476364
MSH3frameshift deletionNM_002439c.1141delAp.K381fsrs587776701
MSH6frameshift insertionNM_001281492c.2916dupTp.T972fsNA
MSH6nonsense mutationNM_001281492c.G726Ap.W242XNA
MSH6splicingNM_001281492g. 48033792 _ 48033795 del TAACNA
MUTYHmissense mutationNM_001128425c.G1187Ap.G396Drs36053993
MUTYHnonsense mutationNM_001128425c.G467Ap.W156Xrs762307622
MUTYHsplicingNM_001128425c.576+1G>CNA
MUTYHsplicingNM_001128425c.934-2A>Grs77542170
MUTYHsplicingNM_001128425c.934-2A>Grs77542170
MUTYHsplicingNM_001128425c.934-2A>Grs77542170
MUTYHsplicingNM_001128425c.934-2A>Grs77542170
MUTYHsplicingNM_001128425c.934-2A>Grs77542170
MUTYHsplicingNM_001128425c.934-2A>Grs77542170
MUTYHsplicingNM_001128425c.934-2A>Grs77542170
MUTYHsplicingNM_001128425c.934-2A>Grs77542170
OGG1nonsense mutationNM_016819c.A974Gp.X325WNA
POLD1splicingNM_002691c.2954-1G>-NA
RAD50frameshift deletionNM_005732c.2157delAp.L719fsNA
RAD50frameshift deletionNM_005732c.536delTp.I179fsNA
RAD50frameshift insertionNM_005732exon13:c.2157dupAp.L719fsrs397507178
RAD51Cframeshift insertionNM_058216c.390dupAp.G130fsrs730881940
RAD51Cnonsense mutationNM_058216c.T833Gp.L278XNA
RAD51CsplicingNM_058216c.905-2A>CNA
RAD51CsplicingNM_058216c.905-2A>CNA
RAD51DsplicingNM_002878c.480+1G>ANA
TP53frameshift deletionNM_000546c.102delCp.P34fsNA
TP53frameshift deletionNM_000546c.121delGp.D41fsNA
TP53frameshift deletionNM_000546c.216delCp.P72fsNA
TP53frameshift deletionNM_000546c.257_272delp.A86fsNA
TP53frameshift deletionNM_000546c.501delGp.Q167fsNA
TP53frameshift deletionNM_000546c.539_549delp.E180fsNA
TP53frameshift insertionNM_000546c.102dupCp.L35fsNA
TP53frameshift insertionNM_000546c.455dupCp.P152fsNA
TP53frameshift insertionNM_000546c.498dupAp.Q167fsNA
TP53frameshift insertionNM_000546c.889dupCp.H297fsNA
TP53missense mutationNM_000546c.A488Gp.Y163Crs148924904
TP53missense mutationNM_000546c.A578Gp.H193Rrs786201838
TP53missense mutationNM_000546c.A659Cp.Y220Srs121912666
TP53missense mutationNM_000546c.A659Gp.Y220Crs121912666
TP53missense mutationNM_000546c.A659Gp.Y220Crs121912666
TP53missense mutationNM_000546c.A736Gp.M246Vrs483352695
TP53missense mutationNM_000546c.A838Gp.R280Grs753660142
TP53missense mutationNM_000546c.C380Tp.S127Frs730881999
TP53missense mutationNM_000546c.C451Tp.P151Srs28934874
TP53missense mutationNM_000546c.C844Tp.R282Wrs28934574
TP53missense mutationNM_000546c.C844Tp.R282Wrs28934574
TP53missense mutationNM_000546c.G412Cp.A138Prs28934875
TP53missense mutationNM_000546c.G524Ap.R175Hrs28934578
TP53missense mutationNM_000546c.G524Ap.R175Hrs28934578
TP53missense mutationNM_000546c.G638Tp.R213Lrs587778720
TP53missense mutationNM_000546c.G730Ap.G244Srs1057519989
TP53missense mutationNM_000546c.G743Ap.R248Qrs11540652
TP53missense mutationNM_000546c.G743Ap.R248Qrs11540652
TP53missense mutationNM_000546c.G818Ap.R273Hrs28934576
TP53missense mutationNM_000546c.G818Ap.R273Hrs28934576
TP53missense mutationNM_000546c.G836Ap.G279Ers1064793881
TP53missense mutationNM_000546c.G856Ap.E286Krs786201059
TP53nonsense mutationNM_000546c.588_589insTGAp.V197delinsXNA
TP53nonsense mutationNM_000546c.912dupTp.K305_R306delinsXNA
TP53nonsense mutationNM_000546c.C430Tp.Q144XNA
TP53nonsense mutationNM_000546c.C499Tp.Q167XNA
TP53nonsense mutationNM_000546c.C574Tp.Q192XNA
TP53nonsense mutationNM_000546c.C586Tp.R196Xrs397516435
TP53nonsense mutationNM_000546c.C637Tp.R213Xrs397516436
TP53nonsense mutationNM_000546c.G272Ap.W91XNA
TP53nonsense mutationNM_000546c.G438Ap.W146XNA
TP53nonsense mutationNM_000546c.G859Tp.E287XNA
TP53nonsense mutationNM_000546c.G880Tp.E294Xrs1057520607
TP53splicingNM_000546c.376-1G>TNA
TP53splicingNM_000546c.672+1G>Ars863224499
TP53splicingNM_000546c.993+2T>GNA
TP53splicingNM_000546c.993+2T>GNA
TP53splicingNM_000546c.993+1G>Trs11575997
TP53splicingNM_000546g.7577493_7577497 del CCTGANA
XRCC4frameshift deletionNM_003401c.810delAp.R270fsNA
XRCC6splicingNM_001469c.589+1G>TNA
Table
Table 4. The correlation of DDR gene mutations with clinical parameters in the epithelial ovarian cancer patients.
Table 4. The correlation of DDR gene mutations with clinical parameters in the epithelial ovarian cancer patients.
GenesHistologyTypeFIGO StageTumor GradeRecurrenceDeath
OSAOEAOCCAIIIEarlyAdvancedLowHighNoYesNoYes
Total172693964104686910329143701029676
HR
Wild type154583660975764902612864908965
89.53%84.06%92.31%93.75%93.27%83.82%92.75%87.38%89.66%89.51%91.43%88.24%92.71%85.53%
Mutation181134711513315612711
10.47%15.94%7.69%6.25%6.73%16.18%7.25%12.62%10.34%10.49%8.57%11.76%7.29%14.47%
p value *0.1540.0480.2590.9810.5020.126
NHEJ
Wild type170683963103676910129141701009674
98.84%98.55%100.00%98.44%99.04%98.53%100.00%98.06%100.00%98.60%100.00%98.04%100.00%97.37%
Mutation21011102020202
1.16%1.45%0.00%1.56%0.96%1.47%0.00%1.94%0.00%1.40%0.00%1.96%0.00%2.63%
p value*0.7420.7610.2440.5220.2390.11
MMR
Wild type161673361956665962413766959170
93.60%97.10%84.62%95.31%91.35%97.06%94.20%93.20%82.76%95.80%94.29%93.14%94.79%92.11%
Mutation112639247564756
6.40%2.90%15.38%4.69%8.65%2.94%5.80%6.80%17.24%4.20%5.71%6.86%5.21%7.89%
p value *0.030.1340.7930.0090.7620.475
BER
Wild type160653758966465952713366949169
93.02%94.20%94.87%90.63%92.31%94.12%94.20%92.23%93.10%93.01%94.29%92.16%94.79%90.79%
Mutation1242684482104857
6.98%5.80%5.13%9.38%7.69%5.88%5.80%7.77%6.90%6.99%5.71%7.84%5.21%9.21%
p value *0.6310.6490.6190.9850.590.306
NER
Wild type167663962102656710029138671009374
97.09%95.65%100.00%96.88%98.08%95.59%97.10%97.09%100.00%96.50%95.71%98.04%96.88%97.37%
Mutation53022323053232
2.91%4.35%0.00%3.13%1.92%4.41%2.90%2.91%0.00%3.50%4.29%1.96%3.13%2.63%
p value *0.430.3420.9960.3070.3730.848
TLS
Wild type171693963103686910229142701019675
99.42%100.00%100.00%98.44%99.04%100.00%100.00%99.03%100.00%99.30%100.00%99.02%100.00%98.68%
Mutation10011001010101
0.58%0.00%0.00%1.56%0.96%0.00%0.00%0.97%0.00%0.70%0.00%0.98%0.00%1.32%
p value *0.4280.4170.4120.6520.4060.26
DSBR
Wild type153573660975664892612764898964
88.95%82.61%92.31%93.75%93.27%82.35%92.75%86.41%89.66%88.81%91.43%87.25%92.71%84.21%
Mutation191234712514316613712
11.05%17.39%7.69%6.25%6.73%17.65%7.25%13.59%10.34%11.19%8.57%12.75%7.29%15.79%
p value *0.0920.0260.1930.8950.3910.077
SSBR
Wild type145603154865959862212359868362
84.30%86.96%79.49%84.38%82.69%86.76%85.51%83.50%75.86%86.01%84.29%84.31%86.46%81.58%
Mutation279810189101772011161314
15.70%13.04%20.51%15.63%17.31%13.24%14.49%16.50%24.14%13.99%15.71%15.69%13.54%18.42%
p value *0.5910.4730.7220.1710.9960.382
CCR
Wild type11929306091286059249557627445
69.19%42.03%76.92%93.75%87.50%41.18%86.96%57.28%82.76%66.43%81.43%60.78%77.08%59.21%
Mutation534094134094454813402231
30.81%57.97%23.08%6.25%12.50%58.82%13.04%42.72%17.24%33.57%18.57%39.22%22.92%40.79%
p value *<0.001<0.001<0.0010.0830.0040.012
DDR
Wild type9421264774205044207447476331
54.65%30.43%66.67%73.44%71.15%29.41%72.46%42.72%68.97%51.75%67.14%46.08%65.63%40.79%
1 gene57357152235144355216412433
mutation33.14%50.72%17.95%23.44%21.15%51.47%20.29%41.75%17.24%36.36%22.86%40.20%25.00%43.42%
2 gene15122131221301541169
mutations8.72%17.39%5.13%1.56%2.88%17.65%2.90%12.62%0.00%10.49%5.71%10.78%6.25%11.84%
3 gene21101111111111
mutations1.16%1.45%2.56%0.00%0.96%1.47%1.45%0.97%3.45%0.70%1.43%0.98%1.04%1.32%
4 gene20202020202020
mutations1.16%0.00%5.13%0.00%1.92%0.00%2.90%0.00%6.90%0.00%2.86%0.00%2.08%0.00%
5 gene10101001100101
Mutations0.58%0.00%2.56%0.00%0.96%0.00%0.00%0.97%3.45%0.00%0.00%0.98%0.00%1.32%
6 gene10011001010101
mutations0.58%0.00%0.00%1.56%0.96%0.00%0.00%0.97%0.00%0.70%0.00%0.98%0.00%1.32%
Total784813173048195996923553345
mutations45.35%69.57%33.33%26.56%28.85%70.59%27.54%57.28%31.03%48.25%32.86%53.92%34.38%59.21%
p value *<0.001<0.001<0.0010.0890.0060.001
Note: BER: base excision repair; CCR: cell cycle regulation; DDR: DNA damage response; DSBR: double-strand break repair; HR: homologous recombination; MMR: mismatch repair; NER: nucleotide excision repair; NHEJ: nonhomologous DNA end joining; OSA: ovarian serous carcinoma; OEA: ovarian endometrioid carcinoma; OCCA: ovarian clear cell carcinoma; SSBR: single-strand break repair; TLS: translesion synthesis. * Pearson’s chi-squared test
Table
Table 5. Cox regression model for the risk factors for recurrence and death in all patients (n = 172).
Table 5. Cox regression model for the risk factors for recurrence and death in all patients (n = 172).
FactorsRecurrenceDeath
UnivariateMultivariateUnivariateMultivariate
nHazard Ratio (95% CI)pHazard Ratio (95% CI)pHazard Ratio (95% CI)pHazard Ratio (95% CI)p
Histology
 OSA691 (reference)1 (reference)1 (reference)1 (reference)
 OEA390.17 (0.08–0.37)<0.0010.42 (0.16–1.12)0.0820.12 (0.04–0.38)<0.0010.45 (0.13–1.55)0.205
 OCCA640.96 (0.64–1.44)0.8351.37 (0.86–2.18)0.188
Type
 I1041 (reference)1 (reference)1 (reference)1 (reference)
 II682.69 (1.81–4.00)<0.0010.77 (0.46–1.28)0.3111.88 (1.19–2.96)0.0070.35 (0.20–0.60)<0.001
FIGO stage
 Early691 (reference)1 (reference)1 (reference)1 (reference)
 Advanced1035.29 (3.16–8.85)<0.0013.08 (1.63–5.80)0.0016.84 (3.28–14.25)<0.0014.82 (2.09–11.09)<0.001
Tumor grade
 Low291 (reference)1 (reference)1 (reference)1 (reference)
 High1435.57 (2.26–13.70)<0.0011.68 (0.55–5.15)0.36617.97 (2.50–129.29)0.0047.38 (0.93–58.28)0.058
Debulking surgery
 Suboptimal601 (reference)1 (reference)1 (reference)1 (reference)
 Optimal1120.28 (0.18–0.41)<0.0010.51 (0.32–0.80)0.0040.26 (0.16–0.41)<0.0010.38 (0.22–0.64)<0.001
HR
 Wild type1541 (reference)1 (reference)
 Mutation181.22 (0.67–2.23)0.5161.15 (0.59–2.25)0.674
NHEJ
 Wild type1701 (reference)1 (reference)
 Mutation22.04 (0.50–8.28)0.3192.52 (0.62–10.32)0.197
MMR
 Wild type1611 (reference)1 (reference)
 Mutation111.31 (0.61–2.83)0.4871.88 (0.81–4.33)0.139
BER
 Wild type1601 (reference)1 (reference)
 Mutation121.32 (0.64–2.71)0.4541.70 (0.78–3.72)0.185
NER
 Wild type1671 (reference)1 (reference)
 Mutation50.58 (0.14–2.36)0.4490.71 (0.18–2.91)0.639
TLS
 Wild type1711 (reference)1 (reference)1 (reference)
 Mutation15.19 (0.71–37.89)0.10433.76 (3.95–289.00)0.0019.57 (1.08–84.83)0.042
DSBR
 Wild type1531 (reference)1 (reference)
 Mutation191.23 (0.69–2.20)0.4881.20 (0.63–2.27)0.584
SSBR
 Wild type1451 (reference)1 (reference)
 Mutation271.10 (0.64–1.87)0.7361.46 (0.82–2.61)0.202
CCR
 Wild type1191 (reference)1 (reference)1 (reference)
 Mutation531.68 (1.12–2.50)0.0110.98 (0.58–1.66)0.9391.54 (0.97–2.45)0.066
DDR
 Wild type941 (reference)1 (reference)1 (reference)1 (reference)
 1 gene mutation571.71 (1.12–2.60)0.0131.18 (0.73–1.91)0.4961.96 (1.20–3.20)0.0071.57 (0.97–2.54)0.062
 ≥2 gene mutations211.52 (0.84–2.76)0.1711.56 (0.78–3.11)0.207
Note: BER: base excision repair; CCR: cell cycle regulation; DDR: DNA damage response; DSBR: double-strand break repair;HR: homologous recombination; MMR: mismatch repair; NER: nucleotide excision repair; NHEJ: nonhomologous DNA end joining; OSA: ovarian serous carcinoma; OEA: ovarian endometrioid carcinoma; OCCA: ovarian clear cell carcinoma; SSBR: single-strand break repair; TLS: translesion synthesis.
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Chiang, Y.-C.; Lin, P.-H.; Lu, T.-P.; Kuo, K.-T.; Tai, Y.-J.; Hsu, H.-C.; Wu, C.-Y.; Lee, C.-Y.; Shen, H.; Chen, C.-A.; et al. A DNA Damage Response Gene Panel for Different Histologic Types of Epithelial Ovarian Carcinomas and Their Outcomes. Biomedicines 2021, 9, 1384. https://doi.org/10.3390/biomedicines9101384

AMA Style

Chiang Y-C, Lin P-H, Lu T-P, Kuo K-T, Tai Y-J, Hsu H-C, Wu C-Y, Lee C-Y, Shen H, Chen C-A, et al. A DNA Damage Response Gene Panel for Different Histologic Types of Epithelial Ovarian Carcinomas and Their Outcomes. Biomedicines. 2021; 9(10):1384. https://doi.org/10.3390/biomedicines9101384

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Chiang, Ying-Cheng, Po-Han Lin, Tzu-Pin Lu, Kuan-Ting Kuo, Yi-Jou Tai, Heng-Cheng Hsu, Chia-Ying Wu, Chia-Yi Lee, Hung Shen, Chi-An Chen, and et al. 2021. "A DNA Damage Response Gene Panel for Different Histologic Types of Epithelial Ovarian Carcinomas and Their Outcomes" Biomedicines 9, no. 10: 1384. https://doi.org/10.3390/biomedicines9101384

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