The PIDDosme in Cancer

Members of the caspase (cystein-dependent aspartate-specific protease) family are considered critical regulators of apoptotic cell death (caspase 3, 6, 7, 8, 9, 10) and inflammation (caspase 1, 4, 5). The physiological role of Caspase-2, the evolutionary most conserved caspase that can become activated in a multi-protein complex containing PIDD (p53-induced protein with a death domain) and the dual adapter molecule RAIDD (receptor-interacting protein (RIP)-associated ICH-1/CED-3 homologous protein with a death domain), dubbed the PIDDosome, however, remains poorly defined. Recent studies suggest a plethora of functions, including roles in apoptosis, cell cycle arrest, as well as DNA repair, some of which may not require PIDDosome formation. Regardless, all these mechanisms are considered tumor suppressive. Consistently, lack of Caspase-2 has been observed in human cancers and Caspase-2 deficiency facilitates oncogene-driven tumor formation in mice. Surprisingly, however, our studies indicate that loss of the upstream activator PIDD delays tumorigenesis in the very same model, in line with proposed Caspase-2-independent functions, suggesting oncogenic properties of PIDD in some settings. However, the molecular mechanism(s) by which PIDDosome components modulate tumorigenesis remain to be defined. The main aims of this revised project are: (A) to explore the impact of oncogenic stress on PIDDosome formation, as well as expression and/or activation of its individual components; (B) to dissect cell autonomous from non-cell autonomous effects of Caspase-2 and PIDD in tumorigenesis; (C) to screen for transcriptome changes in their absence in premalignant cells; (D) to explore the role of individual PIDDosome components in tumorigenesis in different types of blood cancer and (E) to search for novel interactors of individual PIDDosome components and Caspase-2 substrates. We intend to address Aim (A) by conditionally expressing c-Myc (and other selected oncogenes) in model cell lines to assess expression changes and/or activation of PIDDosome components and monitor complex formation. To address Aim (B), we will use c-Myc transgenic fetal liver cells lacking or expressing individual PIDDosome components in adoptive transfer experiments. To address Aim (C), gene chip analysis assessing genome wide expression changes in premalignant E-Myc transgenic B cells lacking or expressing individual PIDDosome components will be performed. To address Aim (D), we plan to explore the role of PIDDosome components in a set of different hematological malignancies and together with clinical partners we will explore their role in human chronic lymphocytic leukemia (CLL). These studies shall be complemented by an unbiased mass- spectrometry based search for interactors of the PIDDosome or its individual components as well as by the identification of Caspase-2 substrates, in Aim (E). From these experiments we anticipate to gain: (i) information on the mechanism(s) that link oncogenic stress and PIDDosome components; (ii) information on cell autonomous vs. non-cell autonomous effects; (iii) insights about the general importance of individual PIDDosome components in oncogene- driven blood cancers in mice and man; (iv) a candidate list of differentially expressed genes and/or interactors as well as putative substrates that may help to define a more detailed mechanism of action and, ideally, novel targets for therapeutic intervention.

Members of the caspase family of proteases are considered critical regulators of two major biological processes, i.e. apoptotic cell death and inflammation. The physiological role of Caspase-2, the evolutionary most conserved caspase that can become activated in a multi-protein complex containing PIDD1, RAIDD, dubbed the PIDDosome, remains poorly defined. Recent studies suggest a plethora of functions, including roles in apoptosis, cell cycle arrest, as well as DNA repair, mechanisms that are considered tumor suppressive. Consistently, lack of Caspase-2 has been observed in human cancer and Caspase-2 deficiency facilitates oncogene-driven tumor formation in mice. Surprisingly, however, our studies also indicate that loss of the upstream activator PIDD1 actually delays tumorigenesis in the very same disease model, in line with its proposed Caspase-2-independent functions, suggesting oncogenic properties, at least in some settings. However, the molecular mechanism(s) by which the PIDDosome components modulate tumorigenesis remained to be defined. The main aims of this project were to explore the impact of oncogenic stress on PIDDosome formation as well as expression and/or activation of its individual components; to dissect cell autonomous from non-cell autonomous effects of Caspase-2 and PIDD1 on tumorigenesis; to explore the role of individual PIDDosome components in tumorigenesis in different blood cancer models and search for novel interactors and/or substrates of the PIDDosome. Overall, we expected to gain a more detailed understanding how the PIDDosome complex affects tumor formation and whether it can be considered a potential drug-target in cancer. Our work yielded the following new insights: 1) In the context of MYC-driven B cell lymphomagenesis the tumor suppressive function of caspase-2 appears to be independent of PIDD1 or RAIDD (Peintner et al., CDD, 2015). 2) Cell death induced by inhibition of checkpoint kinase 1 does not depend on Caspase-2 or the PIDDosome. Yet, CHK1 turns out to be essential for hematopoietic stem cell development in the early embryo, B cell development, function and transformation. (Schuler et. al. Nat Comm. 2017; Schoeler et al. CDD, 2018, Schuler et al. 2019) 3) The PIDDosome is key to activate the p53 tumor suppressor in response to cytokinesis failure and extra centrosomes (Fava et al. Genes & Dev, 2017). 4) The PIDDosome complex controls developmental polyploidization of hepatocytes in the mouse and duringn regeneration in mice and humans. Hence, the PIDDosome consitutes a potential drug target to improve the regenerative capacity of the human liver. (Sladky et al.; submitted; Sladky et al. In preperation).