Omega-3 Docosahexaenoic Fatty Acid Kills Breast Cancer Cells: Unraveling the Complex Pathway that Triggers Cancer Apoptosis Induced by N-3 DHA PolyUnsaturated Fatty Acid

Author: John Hanna

Scheme summarizing the DHA induction of OSGIN1-mediated the onset of apoptosis via the up-regulation of PI3K/Akt/Nrf2 signaling pathway in breast cancer cells

 

Name of paper: Docosahexaenoic acid (DHA) increases the expression of oxidative stress- induced growth inhibitor 1 through the PI3K/Akt/Nrf2 signaling pathway in breast cancer cells

 

Link: https://doi.org/10.1016/j.fct.2017.08.010

 

From our discussions about lipids in BCM 441 over the past few weeks, we started to gain more insights on the different classifications of lipids, especially with regard to poly-unsaturated fatty acids (PUFAs). We learned that these lipids can be categorized based on the position of the first double bond from the methyl end of the fatty acid chain, with each of the omega-3 (N-3) double bond and omega-6 (N-6) double bond being located between the third and fourth carbon, and the sixth and seventh carbon, respectively. Particularly, omega-3 α-linolenic and omega-6 linoleic are essential fatty acids to humans, as they must be acquired from different plant species in order to extend and functionalize them to different metabolites. These fatty acids are extremely beneficial, since they have been linked to improved cardiovascular health, along with decreased risks of depression, Alzheimer’s disease and cancer (Shahidi and Ambigaipalan, 2018). For example, a recent study illustrated that omega-3 eicosapentaenoic and docosahexaenoic acids (EPA and DHA), along with omega-6 gamma-linolenic and arachidonic acids (GLA and AA) all induced apoptosis in the colon cancer cells LoVo and RKO (Zhang et al., 2015). Despite these benefits however, the mechanism in which fatty acids might prevent different cancers from developing has not been fully understood or characterized (Chia-Han et al., 2017). Therefore, in this paper, Chia-Han et al. attempt to both characterize how PUFAs might inhibit the proliferation of breast cancer cells, and develop a full pathway that describes how DHA can lead to the apoptosis of breast cancer tumors.

 

Oxidative stress-induced growth inhibitor 1 OKL38 (or OSGIN1) is a primary tumor inhibitor gene that results in the apoptosis of many carcinoma cells upon its expression (Li et al., 2007). Expression is mediated via the activity of transcription factor nuclear factor E2-related factor (Nrf2), which is a common factor involved in the regulation of many genes that are responsive to oxidative stress in the cell (Li et al., 2007). Under these conditions, Nrf2 translocates from the cytosol to the cell nucleus and forms a heterodimer with small Maf proteins (sMafs) in order to bind to antioxidant response element promoters (AREs) (Kocanova et al., 2007; Li et al., 2008). Studies have also demonstrated that Nrf2 silencing with siRNA significantly reduced stimulated expression of OKL38 in human aortic endothelial cells and thus promoted cancer (Yan et al., 2014). Other players such as extracellular signal regulated kinases (ERKs), c-Jun N-terminal kinases (JNKs), p38 mitogen-activated protein kinases (MAPKs) and phosphoinositide 3-kinase/Akt (PI3K/ Akt) are also involved in Nrf2-mediated gene transactivation, but their roles have not been fully elucidated (Kocanova et al., 2007).

 

Therefore, considering that N-3 and N-6 PUFAs have been linked to the up-regulation of certain apoptosis-related proteins in cancer cells, along with the centrality of OKL38 as a predictive biomarker for tumorigenesis, Chia-Han et al. elected to study the potential of PUFAs to induce the expression of OKL38, and identify a plausible mechanism that results in cancer cell death. Further, considering that breast cancer is one of the most prevalent cancers worldwide, especially among African-American women and other minorities (Desantis et al. 2016), the authors devote their research to MCF-7 and Hs578T human breast cancer cells, and compare the differences to non-tumorigenic mammary epithelial cells (Chia-Han et al., 2017).

 

The authors first assayed the effect of 100 uM AA, LA, GLA, ALA, EPA, and DHA fatty acids on OKL38 expression in both MCF-7 and Hs578T cancer cells, and found that only DHA significantly up-regulated its expression (Chia-Han et al., 2017). They also found that expression was dose-dependent, as lower concentrations of DHA led to decreased expression. They also found that increased concentrations of the other PUFAs had no impact on OKL38 expression. Not only does this distinguish between omega-3 and omega-6 functionalities on OKL38, but also indicates that only omega-3 DHA impacts expression and not omega-3 EPA, which was not expected (Chia-Han et al., 2017). The authors then went on to assess the effect of DHA on the nuclear translocation of Nrf2; and indeed, they demonstrated higher levels of Nrf2 accumulation in the nuclear fraction upon addition of DHA. To verify the role of Nrf2 in DHA-induced OKL38 expression, the authors found that silencing Nrf2 significantly reduced OKL38 expression despite addition of DHA. This suggests that up-regulation of Nrf2 expression and nuclear translocation are critical for the induction of OKL38 by DHA in breast cancer cells (Chia-Han et al., 2017).

 

To further investigate the role of other components in DHA-induced OKL38 expression, the authors also inhibited MAPKs and PI3K/Akt signaling pathways that were shown to up-regulate Nrf2 expression from previous studies (Yang et al., 2013). They found that both DHA-induced OKL38 protein expression and Nrf2 translocation were significantly inhibited by the PI3K inhibitor (Chia-Han et al., 2017). These findings extend the previous results, and suggest that PI3K/Akt signaling also plays a major role in Nrf2-mediated OKL38 expression by DHA (Chia-Han et al., 2017).

 

Considering that the PI3K/Akt signaling pathway is mainly one that is responsive to oxidative stress (Li et al., 2007), Chia-Han et al. elected to test the hypothesis that up-regulation of OKL38 by DHA occurs through the induction of Reactive Oxygen Species (ROS). Indeed, they found an increase in ROS after treating breast cancer cells with DHA. Further, upon adding a ROS scavenger, OKL38 expression was significantly reduced, and translocation of Nrf2 was significantly attenuated, suggesting that DHA deliberately increases ROS in cells, which activates the pathway that leads to tumor suppression.

 

After characterizing the specific function of DHA in inducing breast cancer apoptosis, the authors elected to identify the entire pathway that leads to cancer death. With previous knowledge of proteins that have demonstrated their ability to reduce cancer, such as BCL2 family proteins, p53, caspases, and cytochrome c (Kwon et al., 2008), the authors compared the expression of these proteins in DHA-treated cancer cells to non-DHA treated cells (Chia-Han et al., 2017). They found that Bcl-2 expression decreased while Bax increased in DHA-treated cells. Further, p53 significantly accumulated in the mitochondria, and cytochrome c was notably released from the mitochondria to the cytosol, which initiated cell apoptosis. Therefore, the authors create a scheme that summarizes the apoptosis pathway by DHA induction of OKL38 (refer to cover image). As a summary, the pathway begins with increased ROS induced by DHA, which leads to the activation of PI3K/Akt and translocation of Nrf2 from the cytosol to the nucleus. This then leads to up-regulation of the tumor suppressing protein OKL38, which results in both the increase of the Bax/Bcl-2 ratio and translocation of p53 to the mitochondria. p53 then induces the release of cytochrome c from the mitochondria, which results in the death of the breast cancer cell.

 

Finally, the authors sought to confirm whether the proapoptotic effects of DHA were observed only in breast cancer cells (Chia-Han et al., 2017). They compared MCF-7 breast cancer cells with chemically transformed human mammary epithelial cells (H184) and non-tumorigenic epithelial cells (MCF-10A). They found that treatment with 100 and 200 uM of DHA increased apoptosis in MCF7 cells, but not in MCF-10A. However, as the concentration of DHA increased, more death was observed in MCF-10A. These results suggest that high levels of DHA lead to the death of both breast cancer cells along with normal mammary epithelial cells. Furthermore, accumulation of p53 in the mitochondria and cytochrome c release was significantly higher in MCF7 cells than in both H184 and MCF-10A when induced with 100 uM DHA only. All of these results combined provide evidence for preferential DHA-induced apoptosis in breast cancer cells as opposed to nontumorigenic breast epithelial cells (Chia-Han et al., 2017).

 

This is a tremendous study that elucidates how consuming an omega-3 rich diet, particularly DHA, can potentially prevent from breast cancer. The study has high-impact qualities, considering the characterization of a new apoptosis pathway induced by DHA (and thus ROS ironically), along with the application to both nontumorigenic and cancer breast cells. Other perils from this study are resembled in its potential translational applications in the future to help combat breast cancer. For example, considering that 100 uM DHA triggered apoptosis in cancerous cells but not in normal cells, specific doses can be administered in chemotherapy to help kill cancer cells but cause no harm to the normal cells. Simultaneously, since higher concentrations killed both types of cells, precautions can be established in future clinical therapies in order to provide more efficient and less dangerous treatments. Finally, this study shows that even an excess of omega-3 fats can cause serious health problems. Considering that most studies focus on the harm of excess omega-6 fats and leave out the dangers of excessive omega-3 fat consumption, this study becomes unique in its illustration that DHA can indeed reduce cancer, but also kill normal cells if not consumed in moderation.

 

References:

  1. Tsai, Chia-Han, You-Cheng Shen, Haw-Wen Chen, Kai-Li Liu, Jer-Wei Chang, Pei-Yin Chen, Chen-Yu Lin, Hsien-Tsung Yao, and Chien-Chun Li. 2017. “Docosahexaenoic Acid Increases the Expression of Oxidative Stress-Induced Growth Inhibitor 1 through the PI3K/Akt/Nrf2 Signaling Pathway in Breast Cancer Cells.” Food and Chemical Toxicology 108 (October): 276–88. https://doi.org/10.1016/j.fct.2017.08.010.
  2. Li, Rongsong, Wendy Chen, Rolando Yanes, Sangderk Lee, and Judith A. Berliner. 2007. “OKL38 Is an Oxidative Stress Response Gene Stimulated by Oxidized Phospholipids.” Journal of Lipid Research 48 (3): 709–15. https://doi.org/10.1194/jlr.M600501-JLR200.
  3. Kocanova, Silvia, Esther Buytaert, Jean-Yves Matroule, Jacques Piette, Jakub Golab, Peter de Witte, and Patrizia Agostinis. 2007. “Induction of Heme-Oxygenase 1 Requires the P38MAPK and PI3K Pathways and Suppresses Apoptotic Cell Death Following Hypericin-Mediated Photodynamic Therapy.” Apoptosis 12 (4): 731–41. https://doi.org/10.1007/s10495-006-0016-x.
  4. Li, Wenge, Siwang Yu, Tong Liu, Jung-Hwan Kim, Volker Blank, Hong Li, and A. -N. Tony Kong. 2008. “Heterodimerization with Small Maf Proteins Enhances Nuclear Retention of Nrf2 via Masking the NESzip Motif.” Biochimica et Biophysica Acta (BBA) – Molecular Cell Research 1783 (10): 1847–56. https://doi.org/10.1016/j.bbamcr.2008.05.024.
  5. DeSantis, Carol E., Rebecca L. Siegel, Ann Goding Sauer, Kimberly D. Miller, Stacey A. Fedewa, Kassandra I. Alcaraz, and Ahmedin Jemal. 2016. “Cancer Statistics for African Americans, 2016: Progress and Opportunities in Reducing Racial Disparities.” CA: A Cancer Journal for Clinicians 66 (4): 290–308. https://doi.org/10.3322/caac.21340.
  6. Yan, Xinmin, Sangderk Lee, B. Gabriel Gugiu, Lukasz Koroniak, Michael E. Jung, Judith Berliner, Jinluo Cheng, and Rongsong Li. 2014. “Fatty Acid Epoxyisoprostane E2 Stimulates an Oxidative Stress Response in Endothelial Cells.” Biochemical and Biophysical Research Communications 444 (1): 69–74. https://doi.org/10.1016/j.bbrc.2014.01.016.
  7. Kwon, Jae Im, Gi-Young Kim, Kun Young Park, Chung Ho Ryu, and Yung Hyun Choi. 2008. “Induction of Apoptosis by Linoleic Acid Is Associated with the Modulation of Bcl-2 Family and Fas/FasL System and Activation of Caspases in AGS Human Gastric Adenocarcinoma Cells.” Journal of Medicinal Food 11 (1): 1–8. https://doi.org/10.1089/jmf.2007.073.
  8. Zhang, C., Yu, H., Shen, Y., Ni, X., et al., 2015. Polyunsaturated fatty acids trigger apoptosis of colon cancer cells through a mitochondrial pathway. Arch. Med. Sci. 11, 1081-1094.

 

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