br Furthermore on the basis of ABCG implication
Furthermore, on the basis of ABCG2 implication in ACSL4-over-expressing cell resistance to chemotherapeutic drugs, and given that
this transporter is closely associated to mTOR signaling in other tu-moral lines [44–47], we aimed to establish a connection between mTOR and ABCG2 by assessing ABCG2 protein expression in our breast cancer cell lines. To such end, MCF-7 Tet-Oﬀ/ACSL4 and empty vector LY 379268 were treated with PF4708671 (10 µM), an inhibitor of ribosomal protein S6 kinase 70 kDa polypeptide 1 (p70S6K), a key player in mTOR activation. Interestingly, results showed a sharp decrease in ABCG2 protein expression in MCF-7 Tet-Oﬀ/ACSL4 but no significant varia-tions in MCF-7 Tet-Oﬀ empty vector cells (Fig. 7B). These findings further indicate that ACSL4 participates in cell resistance to cisplatin, doxorubicin and paclitaxel, and that this participation may be partly mediated by its action on ABCG2 through mTOR signaling.
. Identification of drug resistance genes significantly upregulated by ACSL4 in stable human breast cancer cell line MCF7 Tet-Oﬀ/ACSL4 through transcriptome analysis. Data are shown as log2 fold changes in gene expression levels in MCF-7 Tet-Oﬀ/ACSL4 cells relative to MCF-7 Tet-Oﬀ empty vector cells.
Name Gene Symbol Location log2 fold change
ATP-Binding Cassette, Sub-Family C (CFTR/MRP), Member 8 ABCC8 plasma membrane 2.95165 ATP-Binding Cassette, Sub-Family C (CFTR/MRP), Member 4 ABCC4 plasma membrane 2.23521 ATP-Binding Cassette, Sub-Family G (WHITE), Member 2 ABCG2 mitochondrion/nucleus/plasma membrane 1.89082 ATP-Binding Cassette, Sub-Family B (MDR/TAP), Member 8 ABCB8 mitochondrion/nucleus/plasma membrane 1.87866 ATP-Binding Cassette, Sub-Family A (ABC1), Member 7 ABCA7 plasma membrane/endosome/golgi apparatus 1.86503 ATP-Binding Cassette, Sub-Family B (MDR/TAP), Member 10 ABCB10 mitochondrion 1.67862 ATP-Binding Cassette, Sub-Family A (ABC1), Member 2 ABCA2 plasma membrane/endosome/lysosome/vacuole/cytoskeleton 1.66934 ATP-Binding Cassette, Sub-Family F (GCN20), Member 2 ABCF2 plasma membrane/mitochondrion 1.66398 ATP-Binding Cassette, Sub-Family B (MDR/TAP), Member 7 ABCB7 mitochondrion 1.64724 ATP-Binding Cassette, Sub-Family C (CFTR/MRP), Member 5 ABCC5 plasma membrane 1.62059 ATP-Binding Cassette, Sub-Family B (MDR/TAP), Member 9 ABCB9 endoplasmic reticulum/lysosome/vacuole 1.60218
Fig. 4. ABC transporter expression in MCF-7 Tet-Oﬀ cells. Validation of RNA-Seq data through correlation with real-time PCR in MCF-7 Tet-Oﬀ/ACSL4 and MCF-7 Tet-Oﬀ empty vector cells. Data represent log2 ratios of gene expression levels in MCF-7 Tet-Oﬀ/ACSL4 cells relative to MCF-7 Tet-Oﬀ empty vector cells. Spearman’s rank correlation coeﬃcient was 0.500.
Cancer therapeutic approaches usually involve the combination of two or more chemotherapy agents which diﬀer in terms of action and resistance mechanisms [27,28]. The concept of combined chemopre-vention is not new, as most of the world's ancient medicine systems seem to have relied on multiple agents targeting several symptoms at the same time. In particular, combination chemotherapy constitutes a key therapeutic strategy in triple-negative breast cancer and other malignancies. These strategies intend to block multiple intracellular escape pathways essential for tumor survival. Unfortunately, resistance mechanisms to cytotoxic or biological therapies may hinder the ther-apeutic success of anti-cancer drugs, especially in heavily pre-treated patients whose options are rather limited . In this context, this study was undertaken to elucidate the involvement of ACSL4 in anti-cancer drug resistance and the underlying mechanisms.
The most common drugs used for adjuvant and neoadjuvant che-motherapy include anthracyclines, such as doxorubicin and epirubicin, taxanes, such as paclitaxel and docetaxel, 5-fluorouracil (5-FU), cyclo-phosphamide and carboplatin . In addition, recent studies have reported a combination of cisplatin and gemcitabine as a preferred first-line chemotherapy strategy for patients with metastatic triple-negative breast cancer . For these reasons, the drugs used in the current study are among the most common chemotherapeutic agents used to treat breast tumors, although their eﬀects in clinical settings are
thought to be merely additive or, at most, barely synergistic . Regarding ACSL4 participation in cancer resistance mechanisms,
our group and others have established a negative correlation between ACSL4 and estrogen receptor α (ERα) expression, as well as the parti-cipation of ACSL4 in ERα regulation, for instance in metastatic triple-negative cell lines [18,20,21,23]. On the basis of these findings, our group has further demonstrated ACSL4 involvement in resistance me-chanisms hampering anti-estrogenic treatment in in vivo models of triple-negative breast cancer and, most importantly, a significant in-crease in tumor sensitivity to treatment upon the inhibition of ACSL4 . Along the same lines, Wu et al. have reported the participation of ACSL4 in resistance mechanisms undermining anti-androgenic treat-ment in prostate cancer models .