Promoter hypermethylation-mediated down-regulation of LATS1 and LATS2 in human astrocytoma Zheng Jiang a,b,e,1 , Xingang Li b,1 , Jin Hu c, * , Wei Zhou d , Yuquan Jiang b , Gang Li b , Daru Lu e, ** a Laboratory of Neurosurgery, Qilu Hospital of Shandong University, Jinan 250012, China b Department of Neurosurgery, Qilu Hospital of Shandong University, Jinan 250012, China c Department of Neurosurgery, Huashan Hospital, Fudan University, Shanghai 200040, China d Department of Radiotherapeutic Oncology, Shandong Tumor Hospital, Jinan 250117, China e State Key Laboratory of Genetic Engineering, Institute of Genetics, School of Life Sciences, Fudan University, Shanghai 200433, China Received 29 June 2006; accepted 7 September 2006 Available online 17 October 2006 Abstract LATS1 and LATS2 are tumor suppressor genes implicated in the regulation of cell cycle, but their methylation statuses are still unknown in human astrocytoma. Here, we found that the promoter hypermethylation frequencies of LATS1 and LATS1 were 63.66% (56/88) and 71.5% (63/ 88) in 88 astrocytomas by methylation-specific PCR. But no methylation of LATS1 and LATS2 promoter was detected in the 10 normal brain tissues. There was an increased methylation frequency of LATS1 and LATS2 with the malignant development of astrcytoma. By real-time PCR, the mRNA expression of LATS1 or LATS2 was detected significantly decreased in different pathological grade astrocytomas (P < 0.05). And the mRNA levels of LATS1 and LATS2 in astrocytomas with hypermethylation were both significantly (P < 0.01) lower than those without methylation. The methylation of LATS1 and LATS2 was detected in U251 and SHG-44 cell lines, and 5-aza-deoxycytidine could restore their expression to induce cell apoptosis. Our results suggested that LATS1 and LATS2 mRNAwas down-regulated in astrocytoma by hypermethylation of the promoter. The methylation and mRNA expressionof LATS1 and LATS2 may provide useful clues to the development of the diagnostic assays for astrocytoma. Our results also suggested that LATS1 and LATS2 may be a useful target for astrocytoma therapy. # 2006 Elsevier Ireland Ltd and the Japan Neuroscience Society. All rights reserved. Keywords: Astrocytoma; LATS1; LATS2; Methylation; Methylation-specific PCR (MSP) 1. Introduction Diffusely infiltrating astrocytoma is a leading group of the primary central nervous system tumors, accounting for more than 60% of all primary brain tumors (Kleihues and Cavenee, 2000). It may arise aggressively from the normal astrocytes, or evolve stepwise from its benign precursors. Owing to the difficulties with its early diagnosis and surgical removal of all residue diseased tissues, rapid progression, and frequent reoccurrence, the most advanced form of astrocytoma, glioblastoma (WHO grading IV) represents an extremely life-threatening intracranial malignant tumor both inside and outside of China (China Publishing House of Medical Sciences and Technologies, 1998). In China, there are about 35,000 new glioma patients every year, and most of them are astrcytoma patients. It is therefore important to identify new diagnostic approaches and therapeutic targets for this deadly disease. It has emerged that epigenetic events can lead to tumor suppressor gene (TSG) inactivation as an alternative mechan- ism to genetic events, such as gene mutation, deletion or rearrangement. Hypermethylation is a regional event that occurs frequently in GC-rich sequence, called CpG islands and often located within the 5 0 regulatory non-transcribed regions of genes. Aberrant methylation of CpG islands of the promoter www.elsevier.com/locate/neures Neuroscience Research 56 (2006) 450–458 * Corresponding author. ** Corresponding author. Tel.: +86 21 65642799; fax: +86 21 65642799. E-mail addresses: drjiangzheng@hotmail.com (Z. Jiang), lixgang@beelink.com (X. Li), hujindana@sohu.com (J. Hu), zhweiweily@126.com (W. Zhou), yuquanjiang@yahoo.com.cn (Y. Jiang), ligang0710@hotmail.com (G. Li), prof.ludaru@hotmail.com (D. Lu). 1 Equal contributors. 0168-0102/$ – see front matter # 2006 Elsevier Ireland Ltd and the Japan Neuroscience Society. All rights reserved. doi:10.1016/j.neures.2006.09.006