THE ROLE OF AP-1 AND OTHER TRANSCRIPTION FACTORS IN CANCER CAUSE AND PREVENTION

这项研究的总体目标是识别和表征在肿瘤促进和肿瘤进展过程中推动限速步骤的基因调控事件。AP-1转录因子是Jun和Fos家族蛋白的异源二聚体，与某些基因转录启动子上的特定序列结合并驱动其转录。我们1989年的观察(Bernstein和Colburn，Science，1989)发现，转化敏感(P+)而不转化抵抗(P-)的小鼠JB6细胞通过激活AP-1依赖的转录来响应肿瘤促进剂，这表明AP-1的激活可能是从癌前表型发展到肿瘤(肿瘤)表型所必需的。对这一假说的检验表明，药物抑制剂、糖皮质激素和维甲酸以及基因治疗抑制剂显性阴性Jun(TAM67)既能阻断AP-1的激活，又能阻断转化反应(董等人，1994)。这已经扩展到表明特定的维甲酸类化合物在不反式激活视黄酸反应元件(REARE)依赖的基因转录的情况下转录抑制AP-1的活性，也可以防止肿瘤转化(Li等人，癌症资源，1996)。令人惊讶的是，AP-1是由TPA诱导的，而不是由肿瘤坏死因子α诱导的，AP-1对维甲酸的反式抑制很敏感(Li et，Cancer res 1997)。目前的研究旨在确定调节维甲酸对AP-1转录抑制的分子相互作用。由角蛋白14(K14)启动子驱动的显性负Jun突变体(TAM67)在小鼠角质形成细胞系中表达时，抑制了AP-1和NFkB转录因子的活性，并诱导了对Matrigel的侵袭(董等人，Molec)。癌症，1997)提示第二个转录因子NFkB在疾病进展的原因和预防中可能具有重要作用。在人类角质形成细胞进展模型中，AP-1和NFkB活性和DNA结合均呈进行性升高。当转基因K14-TAM67或四环素调控的结构物在更先进的致瘤或锚定非依赖的人类细胞系中表达时，肿瘤细胞的表型被抑制(Li等人，Oncogene，1998和Li等人，提交)。当细胞生长在裸鼠移植床上时，TAM67在小鼠JB6 P+细胞中的表达可使表型逆转为P表型(Strickland等人，癌症。1997年)。获得了表达K14-TAM67转基因的转基因小鼠。在DMBA-TPA启动促进皮肤癌的实验中，这些K14-67小鼠表现出90%的AP-1活性抑制和95%的保护作用，防止皮肤癌变的促进，即防止癌前乳头状瘤的形成(Young等，PNAS，1999)。没有证据表明增殖反应对肿瘤促进剂有毒性或抑制作用。最近，我们发现JB6 P变异体的转化抗性是由于缺乏MAPK激酶ERK1和ERK2，这也限制了AP-1的反式激活(Huang，et al，PNAS，1998)。AP-1反应细胞中显性负性Erk2的表达表明Erk对AP-1的激活和转化反应是必需的(Watts等人，1998)。目前的研究方向是识别激活AP-1所需的ERK依赖的分子事件。最近的观察发现，NFkB无反应性是JB6模型中转化无反应性的一个解释(Hsu等人提交)。因此，以AP-1和NFkB的上调为靶点防止肿瘤的促进和进展的观察已经从小鼠JB6模型扩展到小鼠和人类角质形成细胞进展模型，以及移植和转基因小鼠模型。对限制分子相互作用的新理解正在出现。无论是人角质形成细胞还是表达AP-1抑制因子67的转基因小鼠，都为寻找在肿瘤转化中起关键作用的AP-1靶基因提供了宝贵的机会。-癌症原因，癌症预防，基因调控，转录因子，AP-1，MAPK，NFkappaB，

The overall aim of this research is to identify and characterize gene regulation events that propel rate limiting steps during tumor promotion and tumor progression. The AP-1 transcription factor is a heterodimer of Jun and Fos family proteins that binds to a specific sequence on the transcriptional promoter of certain genes and drives their transcription. Our 1989 observation (Bernstein and Colburn, Science, 1989) that transformation sensitive (P+) but not transformation resistant (P-) mouse JB6 cells responded to tumor promoters by activating AP-1 dependent transcription, suggested that AP-1 activation might be required for progression from preneoplastic to neoplastic(tumor)phenotype. Testing of this hypothesis revealed that the pharmacologic inhibitors, glucocorticoids and retinoids and the gene therapy inhibitor dominant negative jun (TAM67) blocked both AP- 1 activation and transformation response (Dong et al PNAS, 1994). This has been extended to show that specific retinoids that transrepress AP- 1 activity without transactivating retinoic acid response element (RARE) dependent gene transcription, also prevent neoplastic transformation (Li et al., Cancer Res, 1996). Surprisingly TPA induced, but not tumor necrosis factor alpha induced, AP-1 is sensitive to transrepression by retinoid(Li et, Cancer Res 1997). Current research is directed to defining the molecular interactions that regulate AP-1 transrepression by retinoids. The dominant negative jun mutant(TAM67) transgene driven by a keratin 14(K14)promoter, when expressed in a mouse keratinocyte line suppressed both AP-1 and NFkB transcription factor activities as well as induced invasion into matrigel (Dong et al., Molec. Carcinog.,1997)suggesting the possible importance of a second transcription factor NFkB in both the cause and prevention of progression. Both AP-1 and NFkB activities and DNA binding show progressive elevation in a human keratinocyte progression model. When the transgene K14-TAM67 or a tetracycline regulated TAM construct was expressed in the more progressed human cell lines that are tumorigenic or anchorage independent, tumor cell phenotype was suppressed (Li et al., Oncogene, 1998 and Li et al., submitted). Expression of TAM67 in mouse JB6 P+ cells produced phenotypic reversion to P- phenotype when cells were grown on a nude mouse graft bed (Strickland et al., Carcinog. 1997). Transgenic mice expressing the K14-TAM67 transgene have been generated. In DMBA-TPA initiation-promotion skin carcinogenesis experiments these K14-TAM 67 mice show 90% inhibition of AP-1 activation and 95% protection against promotion of skin carcinogenesis, i.e., prevention of premalignant papilloma formation(Young et al., PNAS, 1999). There is no evidence of toxicity or inhibition of the hyperplasia respons to tumor promoter. Recently we have found that the transformation resistance of a JB6 P- variant is due to a shortage of the MAPK kinases Erks 1 and 2 which are also limiting for AP-1 transactivation (Huang, et al, PNAS, 1998). Expression of dominant negative Erk 2 in AP-1 responsive cells has demonstrated the requirement of Erk for AP-1 activation and transformation responses (Watts et al 1998). Current research is directed to identifying Erk dependent molecular events that are required for activating AP-1. Recent observations have identified NFkB non-responsiveness as an explanation for transformation non- responsiveness in the JB6 model(Hsu et al, submitted). Thus the observation that targeting AP-1 and NFkB elevation prevents tumor promotion and progression has been extended from the mouse JB6 model to mouse and human keratinocyte progression models, and to grafting and transgenic mouse models. New understanding of limiting molecular interactions is emerging. Both the human keratinocytes and the transgenic mice expressing AP-1 inhibitor TAM 67 present valuable opportunities to identify AP-1 target genes whose expression is critical to neoplastic transformation. - Cancer Cause, Cancer Prevention, Gene regulation, Transcription Factors, AP-1, MAPK, NFkappaB,