Abstract P1-08-08: Reduction of HER2-associated lipogenic phenotype by docosahexaenoic acid (DHA) induces apoptosis in breast tumor cells harboring HER2 overexpression

Abstract

Abstract The lipogenic phenotype is associated with oncogenic transformation and malignancy in the cancer setting. Our hypothesis is that the oncogenic nature of lipogenesis depends on the expression of HER2 oncogene, and its modulation by docosahexaenoic acid-DHA, might induce apoptosis in breast tumor cells with HER2 overexpression. To evaluate if the lipogenic phenotype might be induced by HER2 overexpression, we used a oncogenic transformation cell model in which, cells were engineered to overexpress HER-2 receptor (HB4aC5.2), but are identical to their parental strain (HB4a) in all other aspects, permitting an specific analysis of enhanced HER-2 effects. Toward this end the lipogenesis profiling was characterized, evaluating several molecular features, including synthesis (FASN), uptake(CD36), transport(FABP4) and storage(DGAT) of FA by RT-PCR and lipogenic regulatory pathways (mTOR, DEPTOR, SREBP1 and PPARγ) in both cells. Lipogenic contribution to lipid rafts formation, which is necessary to HER2 receptor location and activation in the cell membrane, was evaluated by gas chromatography and confocal microscopy. The influence of HER2 overexpression and lipogenic phenotype on proteins activated by HER-2 (AKT, ERK1/2 and FASN) was analyzed by western blot. Next, HB4a and HB4aC5.2 cells were treated, alone or in combination, with DHA, Trastuzumab (anti-HER2), and GW9662 (PPARγ inhibitor) for 72h, and the above experiments were repeated. Cell death was analyzed by flow cytometry and confocal microscopy. Results: In HB4aC5.2 cells, the oncogenic transformation by HER2 overexpression was associated with a lipogenic phenotype, which contributed to increase of lipid rafts formation, activation of survival and proliferation signals, as compared to HB4a normal cells (p<0.001). It is believed that PPARγ is the main regulator of cell lipogenesis. However, our data have shown that mTOR/SREBP pathway, exclusively mTORC1, was decreased concomitantly to increase of DEPTOR gene expression and AKT activation, in HB4aC5.2 cells. Also, the treatment with GW9662 did not change the expression of lipogenic genes, suggesting that lipogenesis seems independent of PPARγ activation, and that HER-2 overexpressing cells may use alternative mechanisms to maintain the lipogenic phenotype. DEPTOR is overexpressed in white adipose tissue and is associated with regulation of lipogenesis. Moreover, DEPTOR overexpression suppresses S6K1 but, by relieving feedback inhibition from mTORC1 to PI3K signaling, activates Akt. In HB4aC5.2 cells, only the DHA treatment decreased the lipogenic phenotype, DEPTOR expression, disrupted lipid raft, inhibited HER-2 signaling, and induced apoptosis (p<0.001). In addition the combined treatment using DHA and Trastuzumab increased cell death. Conclusion: We show that lipogenic phenotype was associated with HER-2 in breast cancer, can be induced early in the oncogenic transformation, and it seems important to promote survival and proliferation. Otherwise, Its modulation by DHA increased death in tumor cells, suggesting this kind of FA may represent a useful tool for controlling the HER2-positive breast cancer. Citation Format: Graziela R Ravacci, Maria M Brentani, William Festuccia, Tharcisio Tortelli, Angela F Waitzberg, Dan L Waitzberg. Reduction of HER2-associated lipogenic phenotype by docosahexaenoic acid (DHA) induces apoptosis in breast tumor cells harboring HER2 overexpression [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P1-08-08.