Abstract 1234: The anticancer drug ABTL0812 induces cancer cell death by impairing Akt/mTORC1 axis and inducing ER stress-mediated cytotoxic autophagy
Authors: Muñoz-Guardiola Pau , Casas Josefina , Megías-Roda Elisabet , Perez-Montoyo Hector , Solé-Sánchez Sonia , Yeste-Velasco Marc , Erazo Tatiana , Diéguez-Martínez Nora , Espinosa-Gil Sergio , Yoldi Guillermo , Muñoz-Pinedo Cristina , Segura Miguel F. , Alfon Jose , Fabriàs Gemma , Velasco Guillermo , Domenech Carles , Lizcano Jose M. 
Published: September 2020

Abstract
ABTL0812 is a first-in-class small molecule with anticancer activity currently in Phase 2a clinical evaluation in patients with advanced endometrial and squamous NSCLC. We have previously described that ABTL0812 induces TRIB3 pseudokinase expression, resulting in inhibition of the Akt-mTORC1 axis and autophagy-mediated cancer cell death. However, classical PI3K/Akt/mTOR inhibitors do not induce an autophagy as strong as ABTL0812 does, therefore we aimed to further elucidate the molecular mechanism responsible for the cytotoxic autophagy which causes ABTL0812 anticancer activity. ABTL0812 induced UPR hallmarks ATF4, CHOP and TRIB3 in vitro in lung, endometrial and pancreatic cancer cell lines, as well as in non-tumor cells. Nevertheless, therapeutic concentrations of ABTL0812 did not induce cytotoxic autophagy in non-tumor cells. Furthermore, genetic or pharmacological inhibition of the UPR resulted in impaired ABTL0812 cytotoxicity in cancer cells. Expression of UPR markers (ATF4, CHOP and TRIB3) in response to ABTL0812 treatment were also validated in xenograft models. In order to uncover the precise molecular mechanism involved in ABTL0812-induced UPR and cytotoxic autophagy we undertook a comprehensive sphingolipidomic analysis, since changes in sphingolipids have been reported to contribute to activation of both UPR and autophagy. ABTL0812 treatment resulted in increased levels of long chain dihydroceramides in cultured cancer cells as well as in vivo. Mechanistically, ABTL0812 impaired desaturase-1 activity (Des-1), the enzyme that introduces a 4,5-trans-double bond in the sphingolipid backbone of dihydroceramides to generate ceramides. Accordingly, in vitro incubation of cancer cells with dihydroceramides resulted in activation of UPR, autophagy and cytotoxicity. Of interest, we observed that tumor cells showed higher Des-1 expression levels than non-tumor cells. Finally, we showed that Des-1 inhibition (GT11) and mTORC1 inhibition (everolimus) collaborate to promote autophagy and cancer cell death, simulating ABTL0812 activity. Furthermore, we have validated the increased expression of CHOP and TRIB3 mRNA levels in blood samples from ABTL0812 treated patients. These biomarkers are currently used as pharmacodynamic biomarkers in the ongoing phase 2 clinical trial in patients with squamous NSCLC and endometrial cancer. To our knowledge, this is the first time that UPR markers are reported to change in human blood in response to any drug treatment. In conclusion, we have shown that ABTL0812 triggers a sustained ER stress and UPR activation mediated by the impairment of Des-1 activity which collaborates with mTORC1 inhibition to induce cytotoxic autophagy in cancer cells, which offers improved anticancer activity over just inhibiting mTORC1.
Citation Format: Pau Muñoz-Guardiola, Josefina Casas, Elisabet Megías-Roda, Hector Perez-Montoyo, Sonia Solé-Sánchez, Marc Yeste-Velasco, Tatiana Erazo, Nora Diéguez-Martínez, Sergio Espinosa-Gil, Guillermo Yoldi, Cristina Muñoz-Pinedo, Miguel F. Segura, Jose Alfon, Gemma Fabriàs, Guillermo Velasco, Carles Domenech, Jose M. Lizcano. The anticancer drug ABTL0812 induces cancer cell death by impairing Akt/mTORC1 axis and inducing ER stress-mediated cytotoxic autophagy [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1234.