Abstract 2751: ASPM, a cell cycle regulated gene and silencing its splicing variant as a molecular target in hepatocellular carcinoma

ASPM gene is the human orthologue of the Drosophila abnormal spindle (asp). As its drosophila orthologue, ASPM protein associates with centrosomes and involves in organizing MTs at the spindle poles in mitosis and meiosis. Homozygous mutation of Drosophila asp protein causes abnormal spindles, frequent polyploid cells and cytokinesis failure. Truncated mice aspm proteins cause mild microcephaly as happen in human, however, cause a massive loss of germ cells in mouse. ASPM participates in spindle organisation, spindle positioning and cytokinesis in dividing cells and interacte with an Angelman syndrome protein UBE3A which is required for proper chromosome segregation. In our previous work, ASPM was shown often overexpressed in human HCC and associated with tumor progression, early tumor recurrence and poor prognosis. In this study, we figured out there were two transcripts (full length/ NM_018136.4 and variant #1/ NM_001206846.1) in human liver tissue. To further elucidate the role of ASPM in liver cancer cells, we used RNAi knockdown to investigate its biological functions in HCC cells. We selected two 21-mers-RNAi oligos named si-482 (at nt5125/exon 18) and si-945 (at nt9977/exon 25) of ASPM mRNA (full length), respectively. Using the RT-PCR detection, the full length ASPM transcript could partially downregulation by both RNAi oligos. However, the ASPM variant#1 transcript only downregulated by the ASPM si-945 RNAi oligos, but could not down-regulate by ASPM si-482 RNAi oligos. Knockdown of ASPM mRNA expression by ASPM si-482 and si-945 RNAi oligos in HuH-7, Ha22T and HCC36 cells and exhibited dramatic decrease HCC cells growth, tumorigenesis and invasion at si-945 but not at si-482 RNAi oligos treatment. We compared the si-482 and si-945 RNAi treatments, than figured out the significance differential expression genes in si-945 RANi treatment but not in si-482 RANi treatment. We identified 197 genes, which were either downregulated (152 genes) or upregulated (45 genes), upon inhibition of ASPM expression. A large fraction of these consequence aberrant expression genes after the ASPM knockdown were involved in tumor-relevant processes, such as signaling transduction, cell cycle regulation, or spindle formation. Numerous cell cycle progression related genes such as NDC80, MIS12, CDC23, TPX2, CEP55, CEP290, DDX21 are identified as the chromosome segregation proteins complexes. However, the chromosome segregation protein, Aurora-A and cyclin B1 also downregulated in protein level, but not in mRNA level. Taken together, our results indicated that ASPM play important roles in cell proliferation, tumorigenesis and cell cycle progression of HCC cells, and suggested a new insight into the mechanism of how ASPM knockdown deregulated TPX2/Aurora-A/Cyclin B1 axis contributes to cell cycle progression of cancer cell and its anti-cancer mechanism in hepatocellular carcinoma. Citation Format: Hung-Wei Pan, Su-Yeh Lin, Tony Wu, Hey-Chi Hsu. ASPM, a cell cycle regulated gene and silencing its splicing variant as a molecular target in hepatocellular carcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2751. doi:10.1158/1538-7445.AM2014-2751