Circular RNA FAT atypical cadherin 1 (circFAT1)/microRNA-525-5p/spindle and kinetochore-associated complex subunit 1 (SKA1) axis regulates oxaliplatin resistance in breast cancer by activating the notch and Wnt signaling pathway

Figures & data

Table 1. Clinicopathologic features of BC patients (n = 66)

Characteristics	Patients
Age (years)
≤50	40
>50	26
Grade
I	6
II	55
III	5
Tumor Size (cm)
≤2	22
>2	44
Node status
Negative (0)	31
Positive (≥1)	35
Estrogen Receptor
Negative	28
Positive	38
Progesterone Receptor
Negative	33
Positive	33
HER 2
Negative	42
Positive	24
Triple-Negative Breast Cancer (TNBC)
Yes	16
No	50

Figure 1. Upregulated circFAT1 is discovered in OX-resistant BC tissues and cells

(A and B) RT-qPCR showed the circFAT1 expression in tissue samples and cells. **P < 0.01, *P < 0.05.

Figure 2. CircFAT1 knockdown decreases resistance of BC cells to OX

(A) RT-qPCR showed the circFAT1 expression levels in BT474/OX and MCF-7/OX cells transfected with sh-circFAT1. (B and C) CCK-8 showed the OX IC₅₀ value in BT474/OX and MCF-7/OX cells treated with increasing OX concentration. (D-G) RT-qPCR and western blotting showed the expression levels of chemoresistance-related genes (MDR1, MRP1, LRP1). *P < 0.05, **P < 0.01.

Figure 3. CircFAT1 inhibition promotes apoptosis and repressed metastatic capabilities of OX-resistant BC cells

(A and B) Flow cytometry showed that the apoptotic ratio of BT474/OX and MCF-7/OX cells transfected with sh-circFAT1 and sh-NC. (C, D) Transwell assay showed the migrated and invaded cells in OX-resistant BC cells transfected with sh-circFAT1 and sh-NC. *P < 0.05, **P < 0.01.

Figure 4. CircFAT1 directly targets miR-525-5p to enhance metastasis and attenuate apoptosis in OX-resistant BC cells

(A) StarBase predicted the binding sites between miR-525-5p and circFAT1. (B) Luciferase reporter assay showed the luciferase activity within miR-525-5p and circFAT1. (C) RT-qPCR showed the miR-525-5p expression in BT474/OX and MCF-7/OX cells compared with parental cells. (D) RT-qPCR showed the miR-525-5p expression in BT474/OX and MCF-7/OX cells transfected with sh-circFAT1. (E) Pearson’s correlation showed the negative correlation between circFAT1 and miR-525-5p in OX-resistant BC samples. (F-H) Flow cytometry and Transwell assays showed the apoptotic and metastatic situation of BT474/OX and MCF-7/OX cells transfected with sh-NC, sh-circFAT1, or sh-circFAT1+ miR-525-5p inhibitor. *P < 0.05, **P < 0.01.

Figure 5. CircFAT1 accelerates metastasis and reduces apoptosis in OX-resistant BC cells via miR-525-5p/SKA1 axis

(A) StarBase website predicted the binding sites between miR-525-5p and SKA1. (B) SKA1 in breast invasive carcinoma (BRCA) tissues, compared with normal tissues according to TCGA database. (C and D) Luciferase reporter assay showed the luciferase activity within miR-525-5p and SKA1. (E-G) Flow cytometry and Transwell assays showed the apoptotic and metastatic situation of BT474/OX and MCF-7/OX cells transfected with NC inhibitor, miR-525-5p inhibitor, or miR-525-5p inhibitor+sh-SKA1. (H and I) RT-qPCR showed the SKA1 expression in BT474/OX and MCF-7/OX cells transfected with si-circFAT1 and/or miR-525-5p inhibitor. (J) Pearson’s correlation showed the negative correlation between SKA1 and miR-525-5p in OX-resistant BC samples. *P < 0.05, **P < 0.01.

Figure 6. SKA1 knockdown hinders notch and Wnt signaling in BC cells resistant to OX

(A) GSEA showed the enrichment plots of gene expression signatures for the Notch/Wnt signaling according to SKA1 expression. (B and C) Western blotting was used to detect the protein expression levels of Notch2, GSK-3β, and β-catenin in BT474/OX cells. **P < 0.01.

Availability of data and materials

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