LOXL2
——具有潛在臨床應用價值的新標志物

魏楓林

(三門峽市中心醫院心胸外科，河南 三門峽 472000)

LOXL2
——具有潛在臨床應用價值的新標志物

魏楓林

(三門峽市中心醫院心胸外科，河南 三門峽 472000)

賴氨酰氧化酶樣蛋白2(Lysyl oxidase-like 2，LOXL2)是賴氨酰氧化酶家族成員之一，其在細胞外主要參與基質膠原蛋白和彈性蛋白的交聯產物的形成，在細胞內主要通過Snail途徑調節上皮細胞-間葉樣細胞轉化(Epithelial-mesenchymal transition，EMT)過程，LOXL2正在作為一種新的治療方法的靶點研究。但是在不同腫瘤中或不同腫瘤亞型中的表達情況差異明顯，需要進一步研究。

賴氨酰氧化酶樣蛋白2；侵襲；轉移

賴氨酰氧化酶樣蛋白2是近年來逐漸受到重視的一種胺氧化酶。它與組織纖維化和腫瘤的侵襲、轉移都有密切的關系，但是在不同類型的腫瘤中，發揮的作用是不同的，甚至是矛盾的。需要我們進一步的深入研究，以了解其中的分子生物學機制，為臨床治療或預測提供一種新的途徑或指標。

1 賴氨酰氧化酶家族

賴氨酰氧化酶樣蛋白2(Lysyl oxidase-like 2，LOXL2)是賴氨酰氧化酶家族成員之一，這個家族具有代表性成員，分別是LOX、LOXL1、LOXL2、LOXL3、LOXL4。它們的-COOH端具有高度保守的序列，這決定了它們的同源性和相似性[1]。然而，LOX家族各亞型的N-端具有更大的差異性，這可能決定了這一組同工酶各自不同的功能角色和組織分布[2]。賴氨酸-酪氨酸輔酶因子和四組氨酸銅結合域是賴氨酰氧化酶家族成員獨有的[3-4]，也是LOX、LOXL2、LOXL4發揮胺氧化酶活性的必要結構[5-8]。賴氨酰氧化酶家族中最具代表性的LOX和LOXL一直被看做是與基質膠原蛋白和彈性蛋白的交聯產物形成密切相關的細胞核外酶[4，9]。然而，Hayashi等[9]和Csiszar等[10]的研究發現，在正常壯年小鼠的各種組織的不同區域細胞的核內和核外都有LOX和LOXL存在，發揮各種對腫瘤的發生和進展密切相關的生物學功能，包括：細胞生長調控[11]、粘附、細胞運動和侵襲[3，12]。

2 LOXL2與腫瘤

1997年，Saito等[13]首次研究發現，相對于粘附性腫瘤細胞株，在各種非粘附腫瘤細胞株中LOXL2表達均下調。在RAS轉化小鼠的纖維母細胞中LOXL2 mRNA的表達減少近60%[14]。還有報道，在頭頸部鱗狀細胞癌(HNSCC)和漿液性卵巢癌中發現LOXL2表達的下調[15-17]。Saux等[18]在1998年將LOXL2基因定位于8p21.2-p21.3，這個區域的染色體丟失現象在多種惡性腫瘤中被發現，包括卵巢癌[19]、頭頸部鱗狀細癌[20]、腸癌[21-22]、食管癌[23-24]和乳腺癌[25-26]。

但是，在新近的研究中卻發現LOXL2的高表達和腫瘤的侵襲行為有關。首先在高致瘤性、轉移性鼠鱗狀細胞癌和梭狀細胞癌中發現LOXL2 mRNA的表達，而同時在無致瘤性的角質細胞株中未發現LOXL2 mRNA的表達[27]。而且同樣的情況也出現在高侵襲性、轉移性乳腺癌細胞株與低侵襲性、轉移性乳腺癌細胞株的對比研究中[3]。有關LOXL2的表達與乳腺癌的臨床腫瘤分級的研究資料也支持上述結果[29]。而且進一步研究發現，原本無侵襲性的MCF-7乳腺癌細胞株被轉染了LOXL2基因后即出現了侵襲性[29]，支持了LOXL2是一種致癌基因的觀點。隨后，有多家分別報道在乳腺癌、腸癌、食管癌、膽管癌、胰腺癌和胃癌細胞和組織中LOXL2的表達升高，而且這種高表達與更差的分化、更高的N分期和臨床TNM分期以及較差的生存期有關[29-33]。

在RAS轉化小鼠的纖維母細胞、頭頸部鱗狀細胞癌、肺腺癌和漿液性卵巢癌中發現LOXL2表達的下調。但是，在乳腺癌、腸癌、食管癌、膽管癌、胰腺癌和胃癌中LOXL2的表達上調，這說明了LOXL2在腫瘤發展過程中的角色是復雜的。而且在Zhan等[34]的報道中，在NSCLC中LOXL2表達的顯著下調，而且和更高的N分期和臨床TNM分期有密切聯系，但這種現象僅僅出現在肺腺癌患者，而在肺鱗癌患者中卻沒有觀察到這種聯系。這更進一步說明了LOXL2的復雜性。

3 LOXL2與EMT

進一步研究發現，在上皮細胞-間葉樣細胞轉化(Epithelial-mesenchymal transition，EMT)過 程 中LOXL2的表達上調，其中包括上皮表型的丟失和間葉細胞的活動性表型的獲得[35]。SNAIL是EMT過程中至關重要的一種轉錄因子[27]。Fong等[29]研究顯示LOXL2能夠與SNAIL相互作用并提高SNAIL蛋白的穩定性，誘導EMT。當SNAIL表達的轉移性癌細胞中LOXL2下調，引起E-cadherin的合成，減少了間葉細胞轉化和促血管生成物質的生成，減低了侵襲性。雖然目前還不確定LOXL2是否通過抑制SNAIL活性的促進侵襲、轉移，但重要的是發現了在EMT過程中LOXL2與NAIL相互作用的重要性，這對于腫瘤的進展，特別是侵襲性和遠處轉移非常重要[29]。LOXL2抗體可以明顯抑制腫瘤的生長和轉移[32]。在上皮細胞株T47D、MCF-7、HCT-116、HCT-15和DLD-1中，LOXL2的表達缺失，而在高侵襲性和轉移性的乳腺癌細胞株MDAMB-231和Hs578T中表達[29]，MDAMB-231和Hs578T具有間葉細胞表型[3]，這與前面的結果是一致的。在一些組織中，例如乳腺和卵巢，早期可能出現LOXL2的表達缺失，但在腫瘤的演化過程中出現LOXL2的再表達，這種現象可能與微環境的變化誘因有關。事實上，MCF-7細胞與纖維母細胞在特定的培養基中共同培養，能夠被促發LOXL2的表達[2]。這和家族中的另一個成員LOX是相近的。LOX已經被證明，根據細胞類型和轉化狀態的不同，可以表現出腫瘤抑制或促腫瘤兩種截然不同的功能[3，36-37]。此外，Peng等[32]研究顯示分泌的LOXL2能夠同時激活Snail/E-cadherin and Src kinase/Focal adhesion kinase(Src/FAK)途徑。然而，LOXL2誘導胃癌細胞侵襲和遠處轉移僅僅通過Src/ FAK途徑。

4 LOXL2的調節

Kaneda等[38]提出，是否象LOX一樣，LOXL2表達調節存在一種外在轉錄調節機制。Fong等[29]用脫甲基因子處理腸癌細胞株，結果發現LOXL2表達的增加。Lind等[39]報道，在腸癌細胞株與原位癌細胞中，LOXL2表達和甲基化率是相同的。然而，但在隨后的腸癌細胞的進展演化中，隨著啟動子的去甲基化，獲得了LOXL2的表達。表達的調控也可能通過腫瘤演化和EMT過程中的轉錄因子完成，如AP1、NFjB和WT1[40-42]。這意味著遺傳和外在環境因素都可能參與了癌癥進展過程中LOXL2基因表達的調節。

綜上所述，LOXL2可能會成為一種預測腫瘤侵襲性和遠處轉移風險的指標，或者成為一種新的治療方法的靶點，但仍需要進一步的研究來證明在體內腫瘤轉移過程中LOXL2的功能作用，探索如何把LOXL2阻斷發展成為一個新的、具有發展潛力的治療策略。

[1]Csiszar K.Lysyl oxidases:A novel multifunctional amine oxidase family[J].Prog NucleicAcid Res Mol Biol,2001,70:1-32.

[2]Molnar J,Fong KS,He QP,et al.Structural and functional diversity of lysyl oxidase and the LOX-like proteins[J].Biochim Biophys Acta,2003,1647(1-2):220-224.

[3]Kirschmann DA,Seftor EA,Fong SFT,et al.A molecular role for lysyl oxidase in breast cancer invasion[J].Cancer Res,2002,62: 4478-4483.

[4]Wang SX,Mure M,Medzihradszky KF,et al.A crosslinked co-factor in lysyl oxidase:Redox function for amino acid side chains[J]. Science,1996,273:1078-1084.

[5]Borel A,Eichenberger D,Farjanel J,et al.Lysyl oxidase-like protein from bovine aorta:Isolation and maturation to an active form by bone morphogenetic protein-1[J].J Biol Chem,2001,276: 48944-48949.

[6]Vadasz Z,Kessler O,Akiri G,et al.Abnormal deposition of collagen around hepatocytes in Wilson's disease is associated with hepatocyte specific expression of lysyl oxidase and lysyl oxidase like protein-2[J].J Hepatol,2005,43(3):499-507.

[7]Ito H,Akiyama H,Iguchi H,et al.Molecular cloning and biological activity of a novel lysyl oxidase-related gene expressed in cartilage [J].J Biol Chem,2001,276:24023-24029.

[8]Kim MS,Kim SS,Jung ST,et al.Expression and purification of enzymatically active forms of the human lysyl oxidase-like protein 4 [J].J Biol Chem,2003,278:52071-52074.

[9]Hayashi K,Fong KS,Mercier F,et al.Comparative immunocytochemical localization of lysyl oxidase(LOX)and the lysyl oxidase-like(LOXL)proteins:changes in the expression of LOXL during development and growth of mouse tissues[J].J Mol Histol, 2004,35(8-9):845-855.

[10]Csiszar K.Lysyl oxidases:A novel multifunctional amine oxidase family[J].Prog NucleicAcid Res Mol Biol,2001,70:1-32.

[11]Butler E,Hardin J,Benson S.The role of lysyl oxidase and collagen crosslinking during sea urchin development[J].Exp Cell Res,1987, 173:174-182.

[12]Payne SL,Fogelgren B,Hess AR,et al.Lysyl oxidase regulates breast cancer cell migration and adhesion through a hydrogen peroxide-mediated mechanism[J].Cancer Res,2005,65:11426-11429.

[13]Saito H,Papaconstantinou H,Sato H,et al.Regulation of a novel gene encoding a lysyl oxidase-related protein in cellular adhesion and senescence[J].J Biol Chem,1997,272:8157-8160.

[14]Zuber J,Tchernitsa OI,Hinzmann B,et al.A genome-wide survey of RAS transformation targets[J].Nat Genet,2000,24:144-152.

[15]Hough CD,Sherman-Baust CA,Pizer ES,et al.Large-scale serialanalysis of gene expression reveals genes differentially expressed in ovarian cancer[J].Cancer Res,2000,60:6281-6287.

[16]Ono K,Tanaka T,Tsunoda T,et al.Identification by cDNA microarray of genes involved in ovarian carcinogenesis[J].Cancer Res, 2000,60:5007-5011.

[17]Rost T,Pyritz V,Rathcke IO,et al.Reduction of LOX-and LOXL2-mRNA expression in head and neck squamous cell carcinomas[J].Anticancer Res,2003,23:1565-1574.

[18]Jourdan-Le Saux C,Le Saux O,Donlon T,et al.The human lysyl oxidase related gene(LOXL2)maps between markers D8S280 and D8S278 on chromosome 8p 21.2-21.3[J].Genomics,1998,51: 301-307.

[19]Lassus H,Laitinen MP,Anttonen M,et al.Comparison of serous and mucinous ovarian carcinomas:Distinct pattern of allelic loss at distal 8p and expression of transcription factor GATA-4[J].Lab Invest,2001,81:517-526.

[20]Bockmuhl U,Schluns K,Kuchler I,et al.Genetic imbalances with impact on survival in head and neck cancer patients[J].Am J Pathol,2000,157:369-375.

[21]Arai T,Akiyama Y,Yamamura A,et al.Allelotype analysis of early colorectal cancers with lymph node metastasis[J].Int J Cancer, 1998,79:418-423.

[22]Lerebours F,Olschwang S,Thuille B,et al.Deletion mapping of the tumor suppressor locus involved in colorectal cancer on chromosome band 8p21[J].Genes Chromosomes Cancer,1999,25: 147-153.

[23]Du Plessis L,Dietzsch E,Van Gele M,et al.Mapping of novel regions of DNA gain and loss by comparative genomic hybridization in esophageal carcinoma in the Black and Colored populations of SouthAfrica[J].Cancer Res,1999,59:1877-1883.

[24]Hu N,Roth MJ,Polymeropolous M,et al.Identification of novel regions of allelic loss from a genomewide scan of esophageal squamous-cell carcinoma in a high-risk Chinese population[J].Genes Chromosomes Cancer,2000,27:217-228.

[25]Yaremko ML,Kutza C,Lyzak J,et al.Loss of heterozygosity from the short arm of chromosome 8 is associated with invasive behavior in breast cancer[J].Genes Chromosomes Cancer,1996,16: 189-195.

[26]Anbazhagan R,Fujii H,Gabrielson E.Allelic loss of chromosomal arm 8p in breast cancer progression[J].Am J Pathol,1998,152: 815-819.

[27]Peinado H,Del Carmen Iglesias-de la Cruz M,Olmeda D,et al.A molec-Genes,Chromosomes&Cancer DOI 10.1002/gccular role for lysyl oxidase-like 2 enzyme in Snail regulation and tumor progression[J].EMBO J,2005,24:3446-3458.

[28]Akiri G,Sabo E,Dafni H,Vadasz Z,et al.Lysyl oxidase-related protein-1 promotes tumor fibrosis and tumor progression in vivo[J]. Cancer Res,2003,63:1657-1666.

[29]Fong SF,Dietzsch E,Fong KS,et al.Lysyl oxidase-like 2 expression is increased in colon and esophageal tumors and associated with less differentiated colon tumors[J].Genes Chromosomes Cancer,2007,46(7):644-655.

[30]Barker HE,Chang J,Cox TR,et al.LOXL2-Mediated Matrix Remodeling in Metastasis and Mammary Gland Involution[J].Cancer Res,2011,71(5):1561-1572.

[31]Ru¨ckert F,Joensson P,Saeger HD,et al.Functional analysis of LOXL2 in pancreatic carcinoma[J].Int J Colorectal Dis,2010,25 (3):303-311.

[32]Peng L,Ran YL,Hu H,et al.Secreted LOXL2 is a novel therapeutic target that promotes gastric cancer metastasis via the Src/FAK pathway[J].Carcinogenesis,2009,30(10):1660-1669.

[33]高應鴻,李天宇,高占峰,等.膽管癌組織中賴氨酰氧化酶樣蛋白-2的表達及其與EMT的關系[J].中華普通外科雜志,2008,23 (10):784-787.

[34]Zhan P,Shen XK,Qian Q,et al.Down-regulation of lysyl oxidase-like 2(LOXL2)is associated with disease progression in lung adenocarcinomas[J].Med Oncol,2012,29:648-655

[35]Kiemer AK,Takeuchi K,Quinlan MP.Identification of genes involved in epithelial mesenchymal transition and tumor progression [J].Oncogene,2001,20:6679-6688.

[36]Erler JT,Bennewith KL,Nicolau M,et al.Lysyl oxidase is essential for hypoxia-induced metastasis[J].Nature,2006,440:1222-1226.

[37]Lucero HA,Kagan HM.Lysyl oxidase:And oxidative enzyme and effector of cell function[J].Cell Mol Life Sci,2006,63:2304-2316. [38]Kaneda A,Kaminishi M,Yanagihara K,et al.Identification of silencing of nine genes in gastic cancers[J].Cancer Res,2002,62: 6645-6650.

[39]Lind GE,Thorstensen L,Lovig T,et al.A CpG island hypermethylation profile of primary colorectal carcinomas and colon cancer cell lines[J].Mol Cancer,2004,3:28-39.

[40]Perez-Pomares JM,PhelpsA,Sedmerova M,et al.Experimental studies on the spatiotemporal expression of WT1 and RALDH2 in the embryonic avian heart:A model for the regulation of myocardial and valvuloseptal development by epicardially derived cells(EPDCs)[J]. Dev Biol,2002,247:307-326.

[41]Eferl R,Wagner EF.AP-1:A double edged sword in tumorigenesis [J].Nat Rev Cancer,2003,3:859-868.

[42]Huber MA,Azoitei N,Baumann B,et al.NF-jB is essential for epithelial-mesenchymal transition and metastasis in a model of breast cancer progression[J].J Clin Invest,2004,114:569-581.

R33

A

1003—6350(2014)15—2258—03

10.3969/j.issn.1003-6350.2014.15.0877

2014-01-08)

魏楓林。E-mail：hhyywfl@163.com