Treating triple negative breast cancer cells with erlotinib plus a select antioxidant overcomes drug resistance by targeting cancer cell heterogeneity

Abstract

Among breast cancer patients, those diagnosed with the triple-negative breast cancer (TNBC) subtype have the worst prog-nosis. TNBC does not express estrogen receptor-alpha, progesterone receptor, or the HER2 oncogene; therefore, TNBC lacks targets for molecularly-guided therapies. The concept that EGFR oncogene inhibitor drugs could be used as targeted treatment against TNBC has been put forth based on estimates that 30–60% of TNBC express high levels of EGFR. However, results from clinical trials testing EGFR inhibitors, alone or in combination with cytotoxic chemotherapy, did not improve patient outcomes. Results herein offer an explanation as to why EGFR inhibitors failed TNBC patients and support how combining a select antioxidant and an EGFR-specific small molecule kinase inhibitor (SMKI) could be an effective, novel therapeutic strategy. Treatment with CAT-SKL—a re-engineered protein form of the antioxidant enzyme catalase—inhibited cancer stem-like cells (CSCs), and treatment with the EGFR-specific SMKI erlotinib inhibited non-CSCs. Thus, combining the antioxidant CAT-SKL with erlotinib targeted both CSCs and bulk cancer cells in cultures of EGFR-expressing TNBC-derived cells. We also report evidence that the mechanism for CAT-SKL inhibition of CSCs may depend on antioxidant-induced downregulation of a short alternative mRNA splicing variant of the methyl-CpG binding domain 2 gene, isoform MBD2c.

Obesity‐induced MBD2_v2 expression promotes tumor‐initiating triple‐negative breast cancer stem cells

“…We designed a set of experiments to examine whether the same regulatory relationship between SRSF2 and MBD2_v2 exists in TNBC cells. First, we observed that expression of SRSF2 is, like MBD2_v2 (Bao et al ., ), subject to antioxidant‐sensitive, ROS‐regulation in TNBC cells. Using MDA‐MB‐468 and SUM149 TNBC cell lines, which expressed similarly abundant endogenous levels of SRSF2, (–)‐epicatechin antioxidant treatment reduced ROS and MBD2_v2 levels (Fig. )…”

“…To more directly test whether increased MBD2_v2 causes increased tumor initiation capacity, we stably overexpressed MBD2_v2 in TNBC cells prior to inoculation. We proceeded to re‐establish, as recently reported using other TNBC lines (Bao et al ., ), that MBD2_v2 overexpression promotes expansion of the CSC fraction in MDA‐MB‐231 TNBC cell cultures using a mammosphere formation assay. Stable overexpression of MBD2_v2 in cells by lentiviral transduction, confirmed by immunoblot and semiquantitative RT‐PCR analysis (Fig.…”

“…), but a possible shortcoming of our study is that we did not attempt to treat DIO mice systemically with (–)‐epicatechin antioxidant in order to affirm that inflammation, ROS specifically, was regulating increased SRSF2 and MBD2_v2 expression in TNBC cell line‐derived tumors as in TNBC cell line cultures (Fig. A) (Bao et al ., ). However, it is well‐documented in the literature that dysfunctional adipose tissue and resident macrophages function as an endocrine organ, producing pro‐inflammatory cytokines that directly act on tumors (van Kruijsdijk et al ., ), and more detailed insights for how the B6.Rag1−/− DIO mouse model system likely parallels human physiology in this regard came to light when we applied genomewide analysis to evaluate the greater impact of DIO on TNBC tumor gene expression.…”

“…We previously identified that ROS-dependent expression of epigenetic reader methyl-CpG-binding domain protein 2 (MBD2), specifically the alternative mRNA splicing variant MBD2_v2, is crucial for maintenance and expansion of self-renewing cancer stem cell-like cells (CSCs) in TNBC cell cultures (Bao et al, 2017). Moreover, in heterogeneous cultures MBD2_v2 expression is contained in the CSC fraction (Bao et al, 2017).…”

Oxidative Stress and Antioxidant Status in Immune Thrombocytopenia

“…However, some other experts considered these therapies are mostly directed towards providing temporary symptomatic relief. Thus many studies have been drifted towards approaches to mitigate this damage by supplementing antioxidant enzymes such as SOD, CAT and glutathione peroxidase [37,38], which could inhibit depolymerization of hyaluron of synovial fluid, inhibiting degradation of neutrophils and proteoglycans in articular cartilage, preventing collagen fragmentation by scavenging reactive oxidants, reducing elastase activity and inhibiting lipid peroxidation [38]. Nevertheless, various reports suggest they have poor pharmacokinetic profile that limits their use.…”