Chemoprevention by black raspberry in oral cancer induced by DB[a,l]P in mice

口腔鳞状细胞癌（OSCC）是头颈部最常见的癌症；在美国，每年有超过45,000例病例和大约11,000例死亡病例。由于缺乏能够反映人类暴露的适当动物模型，在预防和控制OSCC方面的进展受到阻碍。吸烟被认为是口腔癌发生的主要病因。一个有吸引力的动物模型是，由于动物长期暴露于香烟烟雾中存在的化学致癌物而导致的肿瘤，将概括在疾病发展期间在人类OSCC中观察到的生化、分子和细胞改变。我们已经开发了这样一个模型，并建议将其应用于化学预防。先前的研究已经证明，含有5-10%冻干黑覆盆子（BRB）粉末的饮食能够抑制啮齿类动物包括口腔在内的多个器官的化学诱导癌症的发展。7,12-二甲基苯(a)蒽（DMBA）诱导仓鼠颊袋鳞状细胞癌（SCC）。然而，环境中不存在DMBA，仓鼠颊袋模型可能不适用于人类。我们已经证明烟草致癌物二苯并[a，l]芘（DB[a,l]P）可以诱导口腔组织的鳞状细胞癌。其代谢产物（¿）-抗- DB[a，l]PDE是小鼠口腔组织中一种强效的特异性致癌物，DB[a，l]P及其环氧二醇引起的突变谱非常相似，与稳定共价DNA加合物的形成一致（初步结果）。经DB[a，l]P处理的小鼠口腔组织中lacI和p53基因的突变与人类OSCC中观察到的相似。此外，采用LC-MS/MS检测DB[a，l]P和（¿）-抗DB[a，l]PDE对小鼠口腔组织DNA损伤的影响。初步结果显示，饲粮中添加5% BRB粉或0.025%山奈酚（KF）可显著抑制DB[a，l]P- dna加合物在DB[a，l]P小鼠口腔内的形成。我们假设BRB及其活性成分，包括BRB粉末及其花青素富集提取物、原儿茶酸（花青素的主要代谢物）、KF和阿魏酸（覆盆子的成分）将通过多种机制抑制口腔肿瘤的发生，包括：抑制DNA损伤和细胞增殖，调节对癌症进展至关重要的基因，诱导细胞凋亡（目的1），并且在代表疾病不同阶段的人类口腔细胞培养中也将表现出相同的机制作用（目的2）。目标2将扩展和转化我们的发现到未来的临床研究。创新：这项应用是他首次在高度相关的动物模型中研究BRB和相关药物对诱导OSCC的化学预防作用。BRB的化学预防机制也将被研究。相关性/影响：本研究利用相关动物模型对OSCC进行研究，其结果将为未来BRB及其活性化合物的临床化学预防试验制定最有效的策略提供急需的机制见解。

DESCRIPTION (provided by applicant): Oral squamous cell carcinoma (OSCC) is the most common cancer of the head and neck region; in the USA, over 45,000 cases and about 11,000 deaths from the disease occur annually. Progress in the prevention and control of OSCC has been hampered by the lack of appropriate animal models that would reflect human exposure. Tobacco smoking is considered a major etiological factor in the development of oral cancer. An attractive animal model would be one where tumors resulting from chronic exposure of the animal to chemical carcinogens present in cigarette smoke, would recapitulate the biochemical, molecular and cellular alterations observed in human OSCC during the development of the disease. We have developed such a model and propose to apply it to chemoprevention. Previous studies have demonstrated the ability of diets containing 5-10% freeze-dried black raspberry (BRB) powder to inhibit the development of chemically-induced cancers in multiple organ sites in rodents including the oral cavity. 7,12-Dimethylbenz(a)anthracene (DMBA) induced squamous cell carcinomas (SCC) in the hamster cheek pouch. However, DMBA is not present in the environment and the hamster cheek pouch model may not be applicable to humans. We have shown that the tobacco carcinogen, dibenzo[a,l]pyrene (DB[a,l]P) can induce SCC in oral tissues. Its metabolite, (¿)-anti- DB[a,l]PDE is a potent and specific carcinogen in oral tissues in mice and the mutational profiles induced by DB[a,l]P and its diol epoxide are very similar, consistent with the formation of stable covalent DNA adducts (Preliminary Results). Mutations in both lacI and p53 genes in oral tissues of mice treated with DB[a,l]P are similar to those observed in OSCC in humans. In addition, DNA lesions induced by DB[a,l]P and (¿)-anti- DB[a,l]PDE in the oral tissues of mice were detected by LC-MS/MS. Preliminary Results showed 5% BRB powder or 0.025% kaempferol (KF) in diets significantly inhibited DB[a,l]P-DNA adduct formation in the oral cavity of mice treated with DB[a,l]P. We hypothesize that BRB and its active components including BRB powder and its anthocyanins enriched extract, protocatechuic acid (a major metabolite of anthocyanins), KF, and ferulic acid (components of raspberries) will inhibit oral tumorigenesis and do so by multiple mechanisms including: inhibition of DNA damage and cell proliferation, modulation of genes critical to cancer progression, and induction of apoptosis (Aim 1), and will also exhibit the same mechanistic effects in human oral cell cultures representing different stages of the disease (Aim 2). Aim 2 will extend and translate our findings to future clinical studies. Innovation: This application is he first to examine the chemopreventive effects of BRB and related agents on the induction of OSCC in a highly relevant animal model. The mechanisms of chemoprevention by BRB will also be investigated. Relevance/Impact: The results of this study using a relevant animal model for OSCC will provide mechanistic insights that are urgently needed to formulate the most effective strategies for future clinical chemoprevention trials by BRB and its active compounds.