2′‐Hydroxyflavanone inhibits in vitro and in vivo growth of breast cancer cells by targeting RLIP76

Abstract

Consumption of citrus-fruits is associated with reduced incidence of breast cancer (BC), the most common cancer diagnosed in women across the globe. In this study, we investigated the anticancer potential of 2-Hydroxyflavanone (2HF) in BC. 2HF, a citrus-bioflavonoid, has demonstrated anticancer properties in various cancers, but its anticancer role in BC has not been well studied. We investigated the in vitro and in vivo growth inhibitory effects of 2HF in an array of BC lines and in xenograft mouse models of ER-positive and HER2-positive BC cells. Compared to control, 2HF treatment reduced cell viability and suppressed migratory and invasive potential of BC cells, while, no growth inhibitory effects were observed in non-tumorigenic breast epithelial cells. Further, 2HF inhibited the expression of RLIP76, a stress-defensive and anti-apoptotic protein, which is over-expressed in BC cells and simultaneously reduced proliferation of BC cells. Nude mice bearing MCF7 or SKBR3 BC cells xenografts treated with either 2HF or targeting RLIP76 by RLIP76-antisense or RLIP76-antibody treatment had significantly lower tumor-weight as compared to corresponding controls. In addition, Western-blotting and immunohistochemical analysis of tumor tissue from control and treatment group mice showed that 2HF decreased protein expression levels of RLIP76, and the decrease was similar to those seen following RLIP76-antisense treatment. Furthermore, 2HF decreased expression of Ki67, CD31, vimentin, inhibited phosphorylation of Akt and expression of survivin and Bcl2, and increased levels of Bax, E-cadherin, and cleaved-PARP. Therefore, our results indicate that 2HF may suppress BC growth in vitro and in vivo by targeting RLIP76, and may serve as a potential adjuvant treatment in BC patients.

Rlip Depletion Suppresses Growth of Breast Cancer

“…We extensively studied and validated the phosphorothioated Rlip-specific antisense R508 in previous studies for its effects on Rlip depletion, tumor regression, apoptosis, cell cycling, and cell survival [ 7 , 27 , 50 , 51 ]. Previously, we demonstrated that R508-mediated Rlip depletion caused a concentration-dependent decrease in proliferation of the MCF7 cell line in culture [ 52 ]. Locked nucleic-acid-modified antisense oligonucleotides (LNAs) are third generation antisense molecules with improved pharmacological properties.…”

Synergistic efficacy of RLIP inhibition and 2′‐hydroxyflavanone against DMBA‐induced mammary carcinogenesis in SENCAR mice

“…In our recently published studies, we demonstrated the anticancer effects of 2HF in five BC cell lines, covering distinct receptor expression phenotypes: MDA‐MB231 (ER − /PR − /Her2 − ), SKBR3 (ER − /PR − /Her2 + ), MCF7 and T47D (ER + /PR + /Her2 − ), and MDA‐MB361 (ER + /PR − /Her2 + ) . We further demonstrated that the anticancer effect of 2HF was mediated through the inhibition/depletion of RLIP function/expression . In our previous studies on MCF7, MDA‐MB231, and SKBR3 cells, we determined that treating BC cells in vitro with a combination of 2HF and RLIP‐targeting agents reflected a similar response in vivo.…”

“…2HF is a natural phytochemical extracted from oranges that have inhibitory and chemopreventive effects in malignant cell lines . In our recently published studies, we demonstrated the anticancer effects of 2HF in five BC cell lines, covering distinct receptor expression phenotypes: MDA‐MB231 (ER − /PR − /Her2 − ), SKBR3 (ER − /PR − /Her2 + ), MCF7 and T47D (ER + /PR + /Her2 − ), and MDA‐MB361 (ER + /PR − /Her2 + ) . We further demonstrated that the anticancer effect of 2HF was mediated through the inhibition/depletion of RLIP function/expression .…”

“…These findings are consistent with our previous in vitro and in vivo studies that demonstrated significant inhibition of angiogenic and invasive parameters in the human BC MDA‐MB231, SKBR3, and MCF7 cell lines. Our previous studies also demonstrated that the anticancer effects of a mixture of 2HF, RAS, and Rab on BC cell lines both in vitro and in vivo was greater than that of the individual agent …”

“…Further, 2HF inhibited the expression of RLIP, a stress‐defensive and antiapoptotic protein that is overexpressed in BC cells, and simultaneously reduced proliferation of BC cells . The cytotoxic effects of RLIP inhibition via RLIP antibodies or depletion via RAS on BC cells was also investigated using the MTT assay . Here, we used a novel SENCAR mouse model to study the carcinogenic effects of DMBA on the mammary gland.…”