Unsaturated fatty acids maintain cancer cell stemness

Summary

Investigation of the metabolic regulation of cancer stem cells is an emerging field, which offers promising approaches for identifying and targeting recalcitrant stem cell populations. In this issue of Cell Stem Cell Li et al. (2017) indicate that increased lipid desaturation is essential to stem-like characteristics in ovarian cancer cells.

A small but significant percentage of all cancer cells are considered stem-like cells, based on their self-renewal and tumor initiating properties. Cancer stem cells (CSCs) are resistant to standard chemotherapy and are therefore implicated in disease progression, recurrence, and adverse patient outcomes. Evidence now points to a CSC driven hierarchal model of tumor growth, contributing to intra-tumoral heterogeneity and differential responses to chemotherapy. Hence, the need of the hour is to reproducibly identify and characterize cancer stem cell populations as well as to understand the mechanisms that govern their differentiation and pro-metastatic function(s). Several studies have elucidated cellular signaling pathways important to the maintenance of cancer cells stemness; however, the study of metabolic pathways in the regulation of stemness is in its infancy. One of the few indications of CSC metabolic regulation was found in basal-like breast cancer, where the transition of cells along the EMT continuum induces aerobic glycolysis and suppresses ROS production. Switching to glycolysis enhances the cells’ CSC characteristics and allows them to withstand hypoxia in the malignant environment (Dong et al., 2013).

In this issue of Cell Stem Cell, Li et al. (Li et al., 2016) provide new insights into the interdependency of metabolism and stem cell characteristics by comparing the lipid content and composition of ALDH⁺CD133⁺ ovarian CSC with ALDH⁻CD133⁻ cancer cells. In an elegantly performed study, the authors show that ovarian cancer stem cells and cancer cells grown as spheroids have a higher ratio of unsaturated to saturated fatty acids, and that this ratio is essential for the cells to retain stemness. To perform the study, Li et al. use single cell stimulated Raman scattering (SRS) microscopy to compare the lipid content and the levels of unsaturation in the intracellular lipid pools of living single cells (Tipping et al., 2016). By scanning single cells with a femtosecond laser and measuring Raman shifts pertaining to lipid C-H vibrations, the authors show that ovarian CSCs have significantly higher levels of lipid droplets, and that these droplets contain a higher percentage of unsaturated lipids. They then go on to confirm these findings by using primary cancer cells isolated from ascites and grown as spheroids. Given that spheroid culture increases cellular stem-like properties, their data suggest that increases in lipid unsaturation might be a general marker for CSCs in ovarian cancer. In order to determine whether lipid characteristics may be a more generalizable marker for CSC populations, it will be necessary to test multiple CSC populations from different cancers, each isolated using separate cell surface markers. However, the data presented herein strongly suggest that fatty acid desaturation is a metabolic marker of ovarian cancer stem cell populations.

Fatty acids desaturases are a class of enzymes that create double bonds at specific locations in long chain fatty acids. For example palmitic acid (16:0) becomes palmitoleic acid (16:1) with a double bond at the 9^th carbon position. Stearoyl-CoA desaturase-1 (SCD-1), the most abundant desaturase, is expressed in lipogenic tissues and catalyzes the formation of double bonds at the 9^th carbon atom of saturated fatty acids, leading to monounsaturated fatty acids (MUFA). Using chemical inhibitors and molecular approaches (shRNA), the authors identify stearoyl-CoA desaturase-1 (SCD-1) as the enzyme responsible for the increased desaturation in stem cells. They show that blocking SCD-1 significantly reduces the stem cell population, as evidenced by inhibition of several stem cell markers, including aldehyde dehydrogenase activity in vitro. Blocking SCD-1 in ovarian cancer spheroids before injecting them subcutaneously into mice reduces the cells’ capacity to initiate tumors as well as inhibits their proliferation, resulting in very small tumors. Previous reports have shown that SCD-1 is regulated by multiple oncogenic signals and that its expression correlates with cell cycle progression and proliferation (Demoulin et al., 2004), as well as reduced apoptosis and lipid-mediated cytotoxicity (Scaglia and Igal, 2005). In addition, there is strong evidence in the literature that abnormal fatty acid composition, especially an increased MUFA to saturated fatty acid ratio, is present in several cancer types. High levels of SCD-1 and MUFA are associated with an adverse patient prognosis, especially in aggressive breast cancer (Chajes et al., 1999). Li et al. complements these studies by demonstrating, that SCD-1 and MUFA play critical roles in the maintenance of the cancer stem cell phenotype. Moreover, the authors reinforce the view that lipids are not just a static energy depot, but rather lipids are important modulators of the cancer phenotype. This study provides formative evidence that a specific lipid composition and an abundance of lipid droplets promote cancer stemness, corroborating that metabolism affects every aspect of the cancer phenotype.

Finally, using an unbiased approach, the authors identify several stem-cell related pathways affected by SCD-1 inhibition. They demonstrate that NFκB signaling regulates SCD-1, lipid unsaturation, and in consequence, stemness. It is particularly interesting that increased lipid unsaturation and active NFκB signaling upregulate ALDH1A mRNA levels, and that NFκB inhibition abrogates this process. These findings place NFκB at the center of unsaturated lipid-mediated regulation of stemness in ovarian cancer cells. The findings also corroborate previous reports that ovarian cancer cells take up lipids from their adipocyte-rich tumor microenvironment through a FABP4-dependent shuttling mechanism, (Nieman et al., 2011) which is known to affect the NFκB pathway (Makowski et al., 2005).

In summary, this paper is an important contribution to the cancer stem cell literature, as it demonstrates that SCD-1 is critical to maintaining the ovarian CSC phenotype by globally affecting lipogenesis. Chemotherapy is effective for non-stem cells, so SCD-1 inhibition seems to be a promising therapeutic option to specifically target stem cells and prolong patient survival. Several inhibitors of SCD-1 have been developed, mostly for diabetes treatment; however, testing has not gone beyond phase II clinical trials (Zhang et al., 2014). While the trials have not been formally published, deletion of the SCD -1 gene in a mouse model reportedly causes mechanism-based adverse effects such as narrow eye fissures, alopecia, and loss of sebaceous gland function and skin abnormalities (Miyazaki et al., 2001) raising concerns that generalized blocking of SCD-1 may not be a clinically viable approach. However, given the convincing findings of this paper, and the urgent need for effective ovarian cancer treatments especially against stem cells, the identification of an SCD-1 inhibitor that can be clinically tolerated is clearly a worthwhile future pursuit.

Figure 1.

Higher unsaturated lipid profile of ovarian cancer stem cells attributed to increased SCD-1 expression.

Changes in lipid profile were detected using stimulated raman scattering (SRS) microscopy. Ovarian cancer cells grown as spheroids have a higher lipid content, especially unsaturated lipids which scatter monochromatic light differently (left panel) as depicted in the energy level diagram. The authors use this method and show that ovarian cancer stem cells (CSCs) (ALDH⁺/CD133⁺) cells have increased lipid droplets. Growing ovarian CSCs and cell lines as spheroids increase their unsaturated lipid content, which activates NFκB activity, thereby increasing SCD-1 and ALDH1A1 expression. These proteins subsequently increase and maintain stemness of cancer cells, thus inducing a feedforward loop where unsaturated fatty acids increase SCD-1 via NFκB, leading to enhanced unsaturated fatty acid production.