NMI-900

Scalable methods for functionally high-throughput screening of RNA-targeting small molecules are currently limited. Here, an RNA knockdown gene signature and high-throughput sequencing-based high-throughput screening (HTS²) were integrated to identify RNA-targeting compounds. We first generated a gene signature characterizing the knockdown of the long non-coding RNA LINC00973. Then, screening of 8199 compounds by HTS² assay identified that treatments of Hesperadin and GSK1070916 significantly mimic the expression pattern of the LINC00973 knockdown gene signature. Functionally, cell phenotype changes after treatments of these two compounds also mimic the losing function of LINC00973 in multiple types of cancer cells. Mechanistically, the inhibitory action of these two compounds on LINC00973 primarily operates via the AURKB-mediated MAPK signaling pathway, resulting in reduced expression of the transcription factor c-Jun. Consequently, this leads to the suppression of LINC00973 transcription. Moreover, these two compounds significantly inhibit xenograft tumor growth in vivo. Clinically, we further found that breast tumors with high expression of LINC00973 also show relatively high expression of AURKB or JUN, and vice versa. In summary, we established a novel high-throughput screening strategy to identify small molecules capable of targeting RNA, provided two promising compounds targeting LINC00973 and further shed light on the underlying transcriptional upregulation mechanism of LINC00973 within cancer cells.

The impact of the tissue context on tumor growth and drug response in medulloblastoma (MB) is poorly understood. To gain insights into the growth and dissemination behavior of the MB tumor cells under treatment, we combined three-dimensional cell culture screening with ex vivo organotypic cerebellum slice co-culture (OCSC), which allowed the assessment of tumor cell behavior in the tissue context. To identify druggable kinase pathways involved in invasion, we screened a panel of 274 kinase inhibitors and identified aurora kinase B (AURKB) as a potential anti-invasion drug target in MB. We validated tumor suppressive activities of the AURKB inhibitor (AURKBi) Barasertib (AZD1152-HQPA) and the structurally unrelated compound GSK-1070916 in cerebellum slice culture models for SHH, and Grp3 MB. Importantly, AURKBi are tumor suppressive in the tissue context, also in MB tumor cells that are in vitro resistant to the same treatment. We confirmed the requirement of AURKB for tumor growth and expansion in the tissue context through genetic suppression of AURKB by siRNA. We revealed that the combination of AURKBi with the SRC/BCR-ABL inhibitor Dasatinib acts synergistically to repress tumor growth and expansion in the highly invasive MB cell model ONS-76, but not in Grp3 MB cells. We demonstrate that tumor growth in the tissue context is suppressed by pharmacological inhibition of AURKB, comparable to the growth reduction observed after X-ray irradiation, which was used as the positive control. Finally, we show that exposure to µM concentrations of Barasertib does not cause developmental toxicity in fish larvae. In conclusion, we demonstrate that AURKB is essential for MB tumor growth and expansion in the tissue context and the inhibition of AURKB is equally efficient as irradiation in repressing tumor cell growth. In patients younger than three years, pharmacological targeting of AURKB may thus constitute a novel means to overcome radiotherapy limitations.