3.3.3. DDA is an LXR modulator that induces lethal autophagy in cancer cells.
DDA was found to induce cell death and differentiation in mouse and human cancer cells. Further work was done to study the molecular mechanisms involved in DDA cytotoxicity. It was found that DDA induced cytotoxicity in cultured cancer cells in a dose- and time-dependent manner suggesting the implication of a receptor, death was not apoptotic, and required gene expression and protein neosynthesis (Segala et al., 2017), as observed with other ChEH inhibitors (de Medina et al., 2009; Leignadier, Dalenc, Poirot & Silvente-Poirot, 2017; Payre et al., 2008). However, as opposed to other ChEH inhibitors, DDA cytoxicity was not inhibited by anti-oxidants because 5,6-EC do not accumulate and are not second messengers of DDA (de Medina et al., 2009; Leignadier, Dalenc, Poirot & Silvente-Poirot, 2017; Payre et al., 2008; Segala et al., 2017). Studies on the DDA molecular mechanism of cytotoxicity showed that it induced lethal autophagy (Segala et al., 2017) as opposed to other ChEH inhibitors that induced a protective autophagy (Leignadier, Dalenc, Poirot & Silvente-Poirot, 2017). ChEH contributed to autophagy through the accumulation of pro-autophagic Δ8-sterols due the inhibition of its EBP subunit (de Medina, Silvente-Poirot & Poirot, 2009; Silvente-Poirot, Segala, Poirot & Poirot, 2018). DDA was found to be a ligand of LXRα and LXRβ receptors as opposed to other ChEH inhibitors (Segala et al., 2013). Genetic and pharmacological evidences has been given to confirm that LXRβ was required for lethal autophagy. Although DDA was a poor modulator of canonical LXR-dependent genes, it activated via LXRβ the transcription of genes encoding master regulators of lysosome biogenesis and autophagy such as the transcription factor EB (TFEB) (Segala et al., 2017). DDA is, to our knowledge, the first example of an LXR ligand that induces TFEB expression. Such an effect has not been reported to date by other cytotoxic natural LXR ligands belonging to the oxysterols family. It was further established that DDA did not modulate other common nuclear receptors establishing its selectivity to LXRs (Segala et al., 2017).
It was next established that DDA, at cytotoxic doses, actively inhibit the growth of human and mouse tumours implanted into immunocompromized mice with different administration modes, including primary tumours from patients (Segala et al., 2017). Knock down of LXR receptors in cancer cells using small interfering RNA (siRNA) or single hairpin RNA (shRNA) approaches strongly impaired DDA induction of autophagy and its anticancer activities in vitro and in vivo (Segala et al., 2017). This showed that LXRβ was required to control the anticancer activity of DDA. The particular cell death induced by DDA compared to conventional ChEH inhibitors and LXR modulators is probably due to its specific LXRβ-dependent regulation of gene expression and induction of Δ8-sterols accumulation as discussed earlier (Poirot & Silvente-Poirot, 2018). Moreover, this suggests that the control of autophagy might be a specific physiological function of the LXRβ isoform when activated by specific ligands such as DDA. This data shows the importance of LXR in cancer cells as targets for anticancer strategies. Due to the established importance of LXR in the tumour micro-enrironment (Ma & Nelson, 2019) and in the control of the immune response (Fessler, 2016), it will be interesting to determine how autophagy and LXR from immune cells can contribute to the anti-tumour action of DDA observed in immunocompetent mice (de Medina et al., 2013). It would be of interest to determine the impact of combination treatments of DDA with other chemotherapeutic agents acting through different molecular mechanisms, and in particular with drugs that have a limited therapeutic outcome or resistance through the induction of protective autophagy.