RIG-I is expressed in most human cells and plays an important role in the innate immune system (Horvath, Thienpont et al. 2020). As already revealed in functional experiments, its expression can be upregulated by the splicing factor EFTUD2 (Zhu, Xiao et al. 2015). Although EC is known as an “immunogenic” cancer, this is – to the best of our knowledge – the first study examining RIG-I and EFTUD2 in EC. We detected that a high expression of RIG-I correlated with high FIGO- and pT-stages as well as higher grade. Additionally, high expression of RIG-I and EFTUD2 was associated with poor OS- and PFS.
RIG-I is a nucleoid acid receptor in the cytoplasm (Wicherska-Pawłowska, Wróbel et al. 2021). In addition, recent studies showed that RIG-I can be located in the nucleus in some cells (Liu, Lu et al. 2018). The ligands of RIG-I (various RNA molecules) are typically (but not exclusively) parts of the viral RNA (Gack, Shin et al. 2007). After ligand-biding, RIG-I interacts with the MAVS (Breiman, Grandvaux et al. 2005, Matsumiya and Stafforini 2010). This process results in the activation of NFĸB and IRF-3 and leads to the production of Type I IFN, chemokines and other cytokines (Seth, Sun et al. 2005). By these substances, effector T-cells and NK cells are stimulated leading to a tumor-suppressive effect (Elion and Cook 2018). RIG-I can also activate intrinsic and extrinsic apoptosis pathways (Rameshbabu, Labadie et al. 2021). Taking these effects and pathways together, RIG-I as part of the innate immune system has become more and more important over the last years.
In several tumors, RIG-I activation follows the described mechanisms and leads to a tumor-suppressive milieu. Well-studied examples are pancreatic cancer and melanoma (Poeck, Besch et al. 2008, Ellermeier, Wei et al. 2013). RIG-I stimulation by endogenous RNAs enhanced therapy resistance by a STAT-1- dependent pathway and NOTCH signaling (Boelens, Wu et al. 2014). In breast cancer a high RIG-I expression was also described to be associated with poor outcome: RIG-I stimulation by endogenous RNA lead to increased tumor growth and metastasis in vitro (Nabet, Qiu et al. 2017).
This trend also appeared in our results as well as in ovarian cancer. In a retrospective study with 141 cases of ovarian cancer, RIG-I was overexpressed (compared to healthy ovarian tissue) and a high expression correlated with higher tumor grade as well as to poorer outcome (Wolf, Fiegl et al. 2020). Additionally, correlations with interferon beta, PDL-1 and FOXP3 levels were detected in ovarian cancer (Wolf, Fiegl et al. 2020). The immune-suppressive system of PD-L1 was found to be associated with poor survival rates in ovarian cancer as well as in EC (Wieser, Gaugg et al. 2018, Zhang, Liu et al. 2021). The authors suggest that in ovarian cancer the antiviral and tumor-suppressive effects of RIG-I seem to be neutralized by an immune-escape phenomenon in the tumor microenvironment (Wolf, Fiegl et al. 2020). Regarding the tumor microenvironment in EC it is known that regulatory T-cells and their marker FOXP3 are correlated with worse survival rates (de Jong, Leffers et al. 2009, Xi, Jing et al. 2019). By interaction with the tumor microenvironment RIG-I seems to lose its tumor-suppressive effect in ovarian cancer (Wolf, Fiegl et al. 2020). Discovery of mechanism how this protective function has been aborted could help develop a strategies for more effective use of immunotherapies in both EC and OC.
Still, it is important to keep in mind, that our results refer to the expression of RIG-I and not to its activation, similar to the study in ovarian cancer (Wolf, Fiegl et al. 2020). Nevertheless RIG-I remains a potential target in further therapies and agonists as well as antagonists are in development (Rawling, Jagdmann et al. 2020, Onomoto, Onoguchi et al. 2021).
As an upregulation of RIG-I by EFTUD2 was described (Zhu, Xiao et al. 2015), we also investigated the expression of this splicing factor. EFTUD2 seems to be involved in viral infections and miscarriages (Zhu, Xiao et al. 2015b, Vattai et al. 2020). Only limited data is available examining the role of EFTUD2 in cancer and, to the best of our knowledge, this is the first study examining EFTUD2 in EC. We observed a significant correlation between EFTUD2 and survival rates in EC: a high expression was associated with low PFS and OS. EFTUD2 is known to be a modulator of the innate immune system: in colorectal cancer EFTUD2 seems to promote tumor growth, especially in a colitis-associated environment and by modulation of macrophages (Lv, Wang et al. 2019). Lv et. al. demonstrated in a mouse model of colorectal cancer that a knockdown of EFTUD2 resulted in a reduced secretion of proinflammatory cytokines and tumorigenic factors (Lv, Wang et al. 2019). This reduction of inflammation and tumour development was related to an impaired activation of TLR-4-NF-κB signaling cascade in macrophages due to an altered EFTUD2 expression). (Lv, Wang et al. 2019). In hepatic cellular cancer, EFTUD2 was upregulated compared to healthy liver tissue (Lv, Li et al. 2021). In analogy to our results in EC, a negative correlation between survival rates and EFTUD2 expression was shown. Also, a reduced tumor growth in case of EFTUD2 knockdown was detected, suggesting it as an independent prognostic factor for patients with hepatic cellular cancer (Tu, He et al. 2020, Lv, Li et al. 2021).
Furthermore, another limitation of this study needs to be discussed: Although the data regarding FIGO/pT/grade and survival rates are consistent and clear, RIG-I expression did not turn out to be an independent marker for survival. This assumes that underlying mechanisms which are not fully understood play an important role. Further studies are therefore necessary to understand these processes.
In summary, our data show that a high RIG-I expression is correlated with advanced tumor stages and thus subsequently to worse survival rates. EFTUD2 turned out to be an independent marker for survival rates. Therefore RIG – I and EFTUD2 seem to be negative predictors in patients with EC. To examine the underlying mechanisms, further studies are necessary.