Wednesday, April 2, 2025
4/2/2025
Investigating the Immunosuppressive Function of lncRNA CRNDE in Hepatic Tumorigenesis - PROJECT SUMMARY/ABSTRACT Hepatocellular carcinoma (HCC), a malignancy with compromised immune responses in tumor-infiltrating lymphocytes (TILs), is an attractive target for immunotherapy. Studies have demonstrated that the inhibitory immune-checkpoint signaling attributes notably to TIL dysfunction in HCC. Nevertheless, checkpoint blockade treatments exhibited variable outcomes in HCC patients, as indicates the existence of additional mechanisms underlying TIL dysfunction in HCC. Recently, lncRNAs have emerged as important messenger molecules for intercellular communication between malignant cells and infiltrating immunocytes. Our preliminary data revealed that HCC-derived lncRNA CRNDE is transferred into TIL via extracellular vesicle; the transferred CRNDE binds to CypB protein, leading to decreased NFATc activity, TIL dysfunction, and eventually HCC immune evasion and tumor progression. Based on preliminary data, our central hypothesis is that HCC-derived CRNDE promotes tumor development and progression through prompting infiltrating immunocytes (e.g., TILs) dysfunction. The overall objectives of this proposed study are 1) to elucidate the molecular mechanisms by which HCC-derived CRNDE inhibits the anti-tumor immune response of TILs and promotes tumor growth; and 2) to determine the in vivo anti-HCC efficacy of CRNDE inhibition in mouse models. We propose three specific aims to test the central hypothesis and address these highly relevant objectives. In Aim1, the functional impacts of HCC-derived CRNDE on tumor infiltrating immunocytes and tumor progression will be studied by using complementary mouse models. Additionally, ELISpots, FACS, scRNA-seq and other newly-developed methods will be employed to analyze the isolated TILs from mouse models. In Aim2, the specified molecular mechanisms underlying CRNDE-mediated TIL dysfunction will be delineated. Various biochemistry and molecular biology techniques will be employed to scrutinize the detail interaction of CRNDE-CypB, as well as its functional effects on NFATc signaling in TILs. In Aim3, the anti-HCC efficacy of CRNDE-specific LNA gapmer-ASOs and its potential synergic effectiveness with checkpoint blockades will be evaluated by using two complementary mouse HCC models. Given CypB is one of the key targets of immunosuppressants Cyclosporine and FK506/Tacrolimus, this study is highly significant as it is expected to uncover a previously unrecognized mechanism of CypB regulation in the setting of HCC immune evasion. The proposed study is innovative, as it is expected to define a conceptually novel model of HCC immune evasion and to develop a new approach for HCC immunotherapy. At the completion of the proposed study, our expected outcomes are to have defined the mechanisms by which HCC-derived CRNDE suppresses the immune responses of TILs in HCC microenvironments; we also expect to have demonstrated the in vivo anti- HCC efficacy of CRNDE-specific LNA gapmer-ASOs. These results are expected to have an important positive impact on the development of new immunotherapeutic approaches to treat cancer; they will also provide a strong evidence-based proof of principle for further development of ASOs-based therapeutic strategy.
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