Jeon Yoon Kyung, Kim Heejung, Park Soo Oh, Choi Hye Young, Kim Young A, Park Sung-Shin, Kim Ji Eun, Kim Yong Nyun, Kim Chul-Woo
Department of Pathology, Seoul National University College of Medicine, Seoul, Korea.
Lab Invest. 2005 Jul;85(7):874-84. doi: 10.1038/labinvest.3700291.
Extranodal NK/T-cell lymphoma (NKTL), nasal type, is a highly aggressive neoplasm and is strongly associated with Epstein-Barr virus (EBV). In this study, we demonstrate that EBV-positive NKTL cell lines, namely, Hank-1, NK-YS, and NK-L, are resistant to Fas-mediated apoptosis induced by anti-Fas antibodies despite high levels of Fas surface expression and no mutation in the Fas gene. Fas stimulation of Hank-1 and NK-YS cells showed little processing of caspase 8, caspase 3, or bid, although the proximal signaling molecules of the death-inducing signaling complex, namely, Fas, Fas-associated protein with a death domain, caspase 8, and bid were present in these cells. Consistent with previous reports on the hypermethylation of death associated protein (DAP) kinase in NKTLs, the promoter of DAP kinase was methylated and its mRNA not detected in Hank-1 cells. However, the restoration of DAP kinase expression by 5-aza-2'-deoxycytidine did not sensitize Hank-1 to Fas-mediated apoptosis, indicating that DAP kinase deficiency does not contribute to resistance to Fas-mediated apoptosis. Since etoposide-induced apoptosis involved caspase 3 activation in Hank-1 and NK-YS cells, the caspase 3-dependent apoptotic machinery appears to be intact. Interestingly, cotreatment of Hank-1 with cycloheximide, a protein synthesis inhibitor, markedly sensitized cells to Fas-mediated apoptosis along with caspase 8 activation and c-FLIP(L) (cellular FLICE inhibitory protein long form) downregulation. Moreover, immunohistochemistry on paraffin-embedded tissue revealed c-FLIP expression in 39% (14 of 36) of NKTL patients. Taken together, these findings indicate that c-FLIP(L)-mediated resistance to Fas contributes to the development and progression of NKTLs. This study also suggests that agents capable of downregulating c-FLIP(L) could be used to treat NKTL.
结外NK/T细胞淋巴瘤（NKTL），鼻型，是一种高度侵袭性肿瘤，与EB病毒（EBV）密切相关。在本研究中，我们证明EBV阳性的NKTL细胞系，即Hank-1、NK-YS和NK-L，尽管Fas表面表达水平高且Fas基因无突变，但对抗Fas抗体诱导的Fas介导的凋亡具有抗性。Fas刺激Hank-1和NK-YS细胞显示caspase 8、caspase 3或bid几乎没有加工，尽管死亡诱导信号复合物的近端信号分子，即Fas、死亡结构域相关蛋白Fas、caspase 8和bid存在于这些细胞中。与先前关于NKTL中死亡相关蛋白（DAP）激酶高甲基化的报道一致，DAP激酶的启动子在Hank-1细胞中发生甲基化且未检测到其mRNA。然而，用5-氮杂-2'-脱氧胞苷恢复DAP激酶表达并未使Hank-1对Fas介导的凋亡敏感，表明DAP激酶缺乏并不导致对Fas介导的凋亡的抗性。由于依托泊苷诱导的凋亡涉及Hank-1和NK-YS细胞中caspase 3的激活，caspase 3依赖性凋亡机制似乎是完整的。有趣的是，用蛋白质合成抑制剂环己酰亚胺联合处理Hank-1细胞，可使细胞对Fas介导的凋亡明显敏感，同时伴有caspase 8激活和c-FLIP(L)（细胞FLICE抑制蛋白长形式）下调。此外，对石蜡包埋组织的免疫组织化学分析显示，39%（36例中的14例）的NKTL患者中有c-FLIP表达。综上所述，这些发现表明c-FLIP(L)介导的对Fas的抗性有助于NKTL的发生和发展。本研究还表明，能够下调c-FLIP(L)的药物可用于治疗NKTL。
