Tsirulnikov Kirill, Duarte Sergio, Ray Anamika, Datta Nakul, Zarrinpar Ali, Hwang Lin, Faull Kym, Pushkin Alexander, Kurtz Ira
Division of Nephrology, Department of Medicine, D. Geffen School of Medicine, University of California at Los Angeles, Los Angeles, CA, USA.
Dumont-UCLA Transplant Center, D. Geffen School of Medicine, University of California at Los Angeles, Los Angeles, CA, USA.
J Cancer. 2018 Jan 1;9(1):1-12. doi: 10.7150/jca.21747. eCollection 2018.
Ras proteins (HRas, KRas and NRas) are common oncogenes that require membrane association for activation. Previous approaches to block/inhibit Ras membrane association were unsuccessful for cancer treatment in human clinical studies. In the present study we utilized a new approach to decrease Ras membrane association in hepatocellular carcinoma (HCC) cell lines via inhibition of an enzyme aminoacylase 3 (AA3; EC 3.5.1.114). AA3 expression was significantly elevated in the livers of HCC patients and HCC cell lines. Treatment of HepG2 cells with AA3 inhibitors, and HepG2 and HuH7 with AA3 siRNA significantly decreased Ras membrane association and was toxic to these HCC cell lines. AA3 inhibitors also increased the levels of N-acetylfarnesylcysteine (NAFC) and N-acetylgeranylgeranylcysteine (NAGGC) in HepG2 and Huh7 cell lines. We hypothesized that AA3 deacetylates NAFC and NAGGC, and generated farnesylcysteine (FC) and geranylgeranylcysteine (GGC) that are used in HCC cells for the regeneration of farnesylpyrophosphate and geranylgeranylpyrophosphate providing the prenyl (farnesyl or geranylgeranyl) group for Ras prenylation required for Ras membrane association. This was confirmed experimentally where purified human AA3 was capable of efficiently deacetylating NAFC and NAGGC. Our findings suggest that AA3 inhibition may be an effective approach in the therapy of HCC and that elevated AA3 expression in HCC is potentially an important diagnostic marker.
Ras蛋白（HRas、KRas和NRAS）是常见的癌基因，其激活需要与膜结合。以往在人体临床研究中，阻断/抑制Ras与膜结合的方法在癌症治疗中未取得成功。在本研究中，我们采用了一种新方法，通过抑制氨基酰化酶3（AA3；EC 3.5.1.114）来降低肝癌（HCC）细胞系中Ras与膜的结合。AA3在HCC患者肝脏和HCC细胞系中的表达显著升高。用AA3抑制剂处理HepG2细胞，并用AA3 siRNA处理HepG2和HuH7细胞，可显著降低Ras与膜的结合，并对这些HCC细胞系产生毒性。AA3抑制剂还增加了HepG2和Huh7细胞系中N - 乙酰法尼基半胱氨酸（NAFC）和N - 乙酰香叶基香叶基半胱氨酸（NAGGC）的水平。我们推测AA3使NAFC和NAGGC脱乙酰化，并生成法尼基半胱氨酸（FC）和香叶基香叶基半胱氨酸（GGC），这些物质在HCC细胞中用于法尼基焦磷酸和香叶基香叶基焦磷酸的再生，为Ras膜结合所需的Ras异戊二烯化提供异戊二烯（法尼基或香叶基香叶基）基团。这在实验中得到了证实，纯化的人AA3能够有效地使NAFC和NAGGC脱乙酰化。我们的研究结果表明，抑制AA3可能是治疗HCC的有效方法，且HCC中AA3表达升高可能是一个重要的诊断标志物。
