High ATOX1 expression is associated with poor survival of breast cancer patients
Upon survival analysis of all different breast cancer patients in the METABRIC cohort, we discovered that patients with high ATOX1 mRNA levels (10% of patients with highest ATOX1 mRNA levels, i.e., more than a 1.3-fold increase above median) in their primary tumor have poorer survival than those with low ATOX1 mRNA levels (10% of patients with lowest ATOX1 mRNA levels, i.e., more than a 1.3-fold decrease below median) (median DSS is 130 months for low ATOX1 vs. 85 months for high ATOX1, p < 0.001, Fig. 1). Notably, the survival disadvantage is maintained when comparing the 10% of patients with highest ATOX1 mRNA levels (n = 195) vs. all other 90% of patients in the cohort (n = 1709) (Fig. S1). Thus, ATOX1 expression portends a survival disadvantage in breast cancer patients. Since survival is largely coupled to metastasis, high ATOX1 levels may report on increased occurrence of metastasis in those patients.
Follow-up time for the selected cohort (n = 390, i.e., lowest 10% and highest 10% merged) was up to 330 months with a median of 119 months and number of disease-specific events (i.e., breast cancer death) was 137 out of 390 (35.1%) [death by other causes, 87 (22.3%)]. Other clinicopathological parameters that significantly variate between the low and high ATOX1 expression groups are menopausal state (p = 0.024), histological grade (p = 0.044), lymph node status (p < 0.001), human epidermal growth factor receptor 2 (HER2) status (p < 0.001), estrogen receptor (ER) status (p = 0.023) and hormone therapy (p < 0.001) (Table S1). More, larger (> 2 cm) tumors (p = 0.017), histological grade 3 (p = 0.039), positive lymph nodes (p < 0.001), moderate and high cellularity (p < 0.001), and progesterone receptor (PR)-positivity (p = 0.001) correlate with worse disease-specific survival of patients in the selected cohort (Table S2). More, in Fig. S2 we present the number of patients with low (lowest 10%, n = 195) or high (highest 10%, n = 195) ATOX1 mRNA expression as a function of PAM50 molecular subtype and tumor stage. We also included the numbers of patients who received hormone therapy and/or chemotherapy.
High ATOX1 levels relate with worse survival of patients with specific breast cancer subtypes
To obtain more detailed information on the role of ATOX1 in breast cancer, we evaluated the correlation between ATOX1 expression levels and survival outcome for the different PAM50 molecular subtypes, plus the Claudin low molecular subtype (see Fig.S3A, B for KM curves for DSS of patients for different subtypes of breast cancer within the entire cohort (n = 1904) and the selected cohort (n = 390), respectively). The PAM50 subtypes of breast cancer are based upon patterns of gene expression (i.e., 50-gene subtype predictor) and are defined into five groups of Luminal A, Luminal B, HER2-enriched, Basal-like, and Normal-like .
We found a strong correlation between high ATOX1 mRNA levels and reduced DSS for Normal-like (median DSS of 161 months for low ATOX1 vs. 64 months for high ATOX1, p = 0.005), Luminal A (171 vs. 91 months, p = 0.002) and Luminal B (158 vs. 67 months, p = 0.001) breast cancer subtypes. We found no significant correlation between ATOX1 mRNA levels and patient survival for Basal-like (79 vs. 76 months), Claudin low (110 vs. 105 months) and HER2-enriched cancer subtypes (106 vs. 82 months) [Fig. 2, Fig. S4 shows overall survival (OS) KM plots]. This demonstrates that the ATOX1 expression level correlates with patient survival only in specific molecular subtypes of breast cancer.
High ATOX1 mRNA levels relate with worse prognosis for patients at early stages of disease
Next, we investigated ATOX1 expression vs. survival outcome at different stages of breast cancer. While the primary tumor at stages 0 and 1 is localized, at stages 2 and 3 the tumor shows increased regional spread to the lymph nodes . Interestingly, we found the largest correlation between ATOX1 expression levels and disease-specific patient survival for stage 1 breast cancer tumors (median DSS of 164 months for low ATOX1 vs. 100 months for high ATOX1, p = 0.008), followed by a less significant correlation for stage 2 tumors (131 vs. 90 months, p = 0.018) and no correlation for stage 3 tumors (Fig. 3). Since the number of stage 3 cases was very small (n = 18) in the selected cohort, we additionally evaluated the DSS considering the entire cohort (n = 1904) with comparison of 10% highest ATOX1 expression group (n = 195) vs. the lower 90% of the patient cohort (n = 1709) (Fig. S5). Supportive, no significant correlation was detected between ATOX1 expression and survival for stage 3 tumors (48 vs. 59 months). Additionally, considering the entire patient cohort, the difference in survival for stage 2 patients with low vs. high ATOX1 levels became non-significant (p = 0.058). Stages 0 and 4 could not be assessed because there are only few patients with these stages of disease in the METABRIC data set. Thus, the ATOX1 expression level appears to be a determinant of survival only at early stages of breast cancer.