%0 Generic
%A Sorrentino, Antonio
%C Heidelberg
%D 2021
%F heidok:23273
%R 10.11588/heidok.00023273
%T A screening for novel immune modulators identifies  SIK3 kinase as a regulator of tumor resistance to  T cell attack
%U https://archiv.ub.uni-heidelberg.de/volltextserver/23273/
%X Cancer is one of the leading causes of mortality worldwide. An important hallmark of  cancer is its ability to escape immune surveillance by developing several  immunological obstacles. These include a plethora of mechanisms that either dampen  immune cell functionality, or foster tumor cell resistance towards immune attack.  Immunotherapeutic strategies, such as immune checkpoint blockade, have emerged  as promising therapeutic approaches for cancer treatment. However the majority of  tumor patients are refractory to current immune therapeutic tools, emphasizing the  need to identify more key players that could radically improve immunotherapy.  This study aimed to systematically identify novel tumor-associated immune modulators  by performing a high-throughput RNAi screen and subsequently validate novel  candidate genes whose blockade could potentially enhance anti-tumor immune  response in tumor patients. Starting from a pancreatic ductal adenocarcinoma (PDAC)  co-culture model, 2514 genes were knocked down in a luciferase-expressing tumor  cell line using a siRNA library. Subsequently, the transfected tumor cells were cocultured  with HLA-A2.1+- matched tumor infiltrating lymphocytes (TILs). TIL-mediated  cytotoxicity was then assessed by measuring the remaining luciferase intensity of  transfected tumor cells. In order to exclude genes whose knock-down affected cell  viability per se, we cultivated tumor cells with the siRNA library in the absence of TILs.  The primary screening revealed 155 potential candidate genes (hits) whose  downregulation increased T cell-mediated tumor lysis more efficiently than PD-L1  knockdown. The hit-list generated in the primary screen was narrowed down to 108  hits after performing a secondary screen. Among these candidate genes, salt-inducible  kinase 3 (SIK3) was selected for extensive validation analysis, as this protein kinase  is overexpressed in tumor biopsies and its role in immune escape mechanisms has  not been reported so far. SIK3 impairment in tumor cells enhanced T cell mediated  killing in several co-culture models derived from different cancer entities. We showed  that SIK3 sustained intrinsic tumor resistance to immune cell attack, rather than  modulating T cell functionality. We observed that T cells expressed TNF-α upon coculture  with tumor cells, and that this cytokine elicited tumor cell growth in SIK3-  proficinent cells. On the contrary, SIK3 depletion sensitized tumor cells towards TNFPage  VIII  α-induced apoptosis by regulating NF-κB activation via HDAC4. To prove the  translational relevance of SIK3 blockade for cancer immunotherapy, we used a small  molecule compound which recapitulated the effect of SIK3 genetic depletion.  Additionally, stable knockdown of SIK3 in tumor cells resulted in retardation of tumor  growth after adoptive cell transfer of TILs in a xenograft mouse model.  This study describes a robust method for a comprehensive identification of novel  immune modulators in solid tumors. Furthermore, this work provides the rationale of  SIK3 inhibition as a novel therapeutic strategy for cancer treatment.