br For The Cancer Genome Atlas TCGA cohort the
For The Cancer Genome Atlas (TCGA) cohort, the KMT2C somatic mutation profiles and clinicopathological information of breast cancer patients were downloaded from the website http://genome-cancer. ucsc.edu. A total of 981 patients with breast cancer were eligible for this study. For the Molecular Taxonomy of Breast Cancer International Consortium (METABRIC) cohort , 1454 patients with breast cancer with complete KMT2C somatic mutation profiles and clin-icopathological characteristics were collected for this study (http:// www.cbioportal.org). Biomedicine & Pharmacotherapy 116 (2019) 108997
2.2. Tissue DNA extraction
DNA was extracted from formalin-fixed paraffin-embedded tissue sections using QIAamp (Qiagen, CA, USA) according to the manu-facturer’s instructions. The DNA concentration was measured using the Qubit dsDNA Assay Kit (Life Technologies, CA, USA).
2.3. NGS library preparation and capture-based targeted DNA sequencing
The sequencing results of the primary tumor tissue samples were obtained. Procedures including tissue sample preparation, NGS process, and data processing were conducted according to a previous study .
2.4. Statistical analysis
All data were analyzed using SPSS 22.0 software (Chicago, IL, USA). Pearson’s chi-squared (χ2) test, Fisher’s exact test, and the Kruskal–Wallis test were used to test the associations between different variables. Kaplan–Meier survival analysis and the Cox proportional hazard models were used to evaluate the relationship between KMT2C mutation and overall survival (OS) in the TCGA and METABRIC co-horts. All data are presented as mean ± SD. The results with p < 0.05 were considered statistically significant.
3.1. Clinicopathological features of the GDPH, TCGA, and METABRIC cohorts
The clinicopathological features of patients in the three databases were significantly different, including age of onset (p < 0.001), me-nopausal status (p < 0.001), tumor size (p < 0.001), ZVADFMK status
3.2. KMT2C mutation spectrum of patients in the GDPH, TCGA, and METABRIC cohorts
Clinicopathologic characteristics of breast cancer patients in GDPH, TCGA, and METABRIC cohorts.
TCGA, The Cancer Genome Atlas; METABRIC, Molecular Taxonomy of Breast Cancer International Consortium; GDPH, Guangdong Provincial People's Hospital; ER, estrogen receptor; PR, progesterone receptor; HR, Hormone receptor; HER2, human epidermal growth factor receptor 2; Ki-67, Ki-67 protein; AR, androgen receptor; MT, mutant; NA, not applicable; WT, wild-type; pTNM, pathological TNM.
Fig. 1. Distribution of KMT2C mutation types in GDPH, TCGA and METABRIC cohorts.
domains of KMT2C in the METABRIC cohort are shown in Fig. 2C. These results showed that truncated and missense mutations were the main mutation types of the KMT2C gene, accounting for approximately 90% in the three cohorts. Biomedicine & Pharmacotherapy 116 (2019) 108997
3.3. Relationship between KMT2C mutation and clinicopathological features
The patients in the three databases were divided into the KMT2C mutant and wild-type groups, respectively, and the relationship be-tween the clinicopathological characteristics of patients with breast cancer and KMT2C mutation status was compared and analyzed. In the three groups, we found that female patients over 50 years of age were more prone to KMT2C mutation (GDPH: p = 0.007; TCGA: p = 0.005; METABRIC: p = 0.015). The KMT2C mutation rate in HR+/HER2-breast cancer was higher than that in the other subtypes (GDPH: p = 0.047; TCGA: p = 0.032; METABRIC: p = 0.046). Additionally, KMT2C mutations were observed in 30.8% (4/13) of patients in the GDPH cohort with invasive lobular breast cancer (ILC), but there was no significant association with ILC and KMT2C mutation status in the other two cohorts (TCGA: p = 0.813; METABRIC: p = 0.269). The relation-ship between KMT2C mutation and clinicopathological features is summarized in Table 4.
Fig. 2. Diagram of KMT2C domains with the mutation location identified in GDPH, TCGA and METABRIC cohorts. (A) GDPH cohort; (B) TCGA cohort; (C) MET-ABRIC cohort.
Mutation domains of KMT2C of reported studies were detected in GDPH cohort.
No. Exon# Mutation Mutation types Mutation ID Tumor types Refs.
2 14 p.E728* Nonsense COSM1449460 Biliary tract Carcinoma; Nasoph aryngeal carcinoma; Colon adenocarcinoma 
Endometrioid carcinoma; Skin basal cell carcinoma; Stomach intestinal adenocarcinoma;