Genomic Feature Analysis of Hereditary Female Cancers based on Next Generation Sequencing

基於二代測序數據的女性遺傳癌症基因特征分析

Student thesis: Doctoral Thesis

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Award date18 Aug 2023

Abstract

In this thesis, we studied Genomic Feature Analysis of Hereditary Female Cancers based on Next Generation Sequencing. Breast cancer and ovarian cancer are two highly threatening malignancies affecting women. Breast cancer has the highest incidence among women, while ovarian cancer ranks third in incidence but carries the highest mortality rate. These cancers are closely linked to genetic factors, with approximately 10%-20% of cases being associated with hereditary predisposing gene mutations. Extensive research has identified over 20 genes associated with hereditary breast and ovarian cancer, as well as more than 100 genes associated with other hereditary cancers. Notably, the BRCA1 and BRCA2 genes have been extensively studied, and advancements in next-generation sequencing technology have facilitated multi-gene research.

Mutations in BRCA1, BRCA2, and related homologous recombination (HR) pathway genes not only increase the risk of breast cancer and ovarian cancer but also impact treatment strategies and prognosis for these cancers. Current research primarily focuses on the frequency of susceptibility gene mutations in different populations, the association of mutations with chemotherapeutic and targeted treatment drug sensitivity, and the survival differences between mutation carriers and non-carriers. However, variations in race, target populations, and interpretation may lead to different conclusions.

This thesis aims to investigate the association between sets of hereditary predisposing genes and clinical characteristics in breast and ovarian cancer. Additionally, it seeks to elucidate the genomic profiling and potential clinical applications of susceptibility gene detection in different populations. To achieve these objectives, we conducted multi-gene panel studies on breast and ovarian cancer in Guatemala and China, and collaborated with 41 laboratories to evaluate their interpretation ability of predisposing genes.

In the first study, target sequencing of epithelial ovarian cancer (EOC) was employed to detect the status of BRCA1/2 and genes in the homologous recombination repair (HRR) pathway. A total of 229 EOC patients underwent ovarian cancer multi-gene panel sequencing with 25 genes, while 141 patients received pan-cancer panel sequencing with 508 genes. The findings revealed that carriers of BRCA2, BRCA1/2, and HRR mutations exhibited longer overall survival (OS) and progression-free survival (PFS) compared to non-carriers in univariate or multivariate analysis. Furthermore, classifying variants of uncertain significance (VUS) as deleterious mutations resulted in prolonged OS and PFS for mutation carriers, and non-BRCA HRR mutation carriers displayed significantly longer PFS. Regarding platinum-based chemotherapy, germline and somatic pathogenic variants in BRCA1, BRCA1/2, and HRR-related genes were associated with better treatment response. Moreover, carriers of germline and somatic pathogenic variants in BRCA1, BRCA1/2, and HRR mutations exhibited significant sensitivity to the PARP inhibitor Niraparib.

The second study aimed to investigate the prevalence, clinical significance, and impact on prognosis of germline mutations in 21 cancer-predisposing genes in EOC. We conducted a comprehensive analysis of mutation spectra in a multicenter retrospective cohort study involving Chinese patients with EOC. The mutation prevalence in EOC was calculated and compared to two other large cohorts. The association between clinical characteristics, survival, and mutation status was assessed. Out of 961 subjects, 171 (17.8%), 61 (6.3%), and 26 (2.7%) had deleterious variants in BRCA1, BRCA2, and other HRR genes, respectively. Patients carrying BRCA1 and other HRR gene variants exhibited a higher likelihood of achieving a platinum-sensitive response. Survival outcomes between patients with BRCA1/2 LP+ and BRCA1/2 VUS were similar in univariate analysis. After adjusting for age, histology, and FIGO stage, BRCA1/2 VUS+ variants were associated with improved overall survival (adjusted HR: 0.57, 95% CI: 0.35-0.90, p = 0.017), and BRCA2 VUS+ variants were associated with improved progression-free survival (adjusted HR: 0.59, 95% CI: 0.38-0.92, p = 0.019). These findings highlight the positive impact of BRCA1/2 and other HRR gene mutations on platinum responsiveness. Additionally, BRCA1/2 VUS+ variants significantly extended OS and PFS and should not be overlooked.

The third study focused on the role of mutations in hereditary breast cancer genes in cancer risk. We sequenced cancer susceptibility genes in 664 unselected breast cancer cases from Guatemala. The most frequently mutated genes were BRCA1 (37/664, 5.6%), followed by BRCA2 (15/664, 2.3%), PALB2 (5/664, 0.8%), and TP53 (5/664, 0.8%). Pathogenic variants were also identified in moderate penetrance genes such as ATM, BARD1, CHEK2, and MSH6. The high ratio of BRCA1/BRCA2 mutations can be attributed to two potential founder mutations: the BRCA1 c.212+1G>A splice mutation (15 cases) and the BRCA1 c.799delT mutation (9 cases). Cases with pathogenic mutations had a significantly earlier age at diagnosis (45 vs 51 years, p < 0.001), a higher likelihood of premenopausal diagnosis, and a higher percentage had relatives with any cancer (51% vs 37%, p = 0.038) or breast cancer (33% vs 15%, p < 0.001).

The fourth study evaluated the performance of interpretation systems in assessing the capacities of BRCA1/2 interpretation across different laboratories in China. The evaluation was conducted among 41 laboratories, and their performance was categorized into five levels. Only Level A, which allowed for a 0.3% error rate of clinically relevant misinterpretation, was considered qualified. Among the laboratories, 26 out of 41 (63%) met the qualified standard, while 7 laboratories were classified as levels D and E, indicating significant mistakes and systemic issues in variant interpretation. Due to the strict quality requirements, the interpretation of several variants was amended, which had a significant impact on the quality rate. If the incorrect recommended interpretations were not corrected, the number of qualified laboratories would decrease from 26 to 17.

In summary, our findings demonstrate that in ovarian cancer, BRCA2 and BRCA1/2 mutation carriers exhibit longer OS and PFS than non-carriers. Considering VUS improves the robustness of the survival benefit. Notably, BRCA1, BRCA1/2, and HRR-related genes can be used to predict drug sensitivity. The combination of germline and somatic variants enhances performance. We also observed hereditary breast cancer mutations in Guatemalan women, who were more likely to be diagnosed at an early age and have a family history of cancer. These findings suggest the utility of genetic testing in breast cancer patients and those at high risk as part of a strategy to reduce breast cancer mortality in Guatemala. Furthermore, the evaluation system provides a potential approach for standardizing variant interpretation and reducing discordance between different laboratories. A well-designed interpretation ability evaluation is crucial for assessing laboratory proficiency in real-world clinical settings.