DEVELOPMENT OF A STOCHASTIC MOUSE TO STUDY BREAST CANCER

尽管70%死于乳腺癌的患者有溶骨性骨转移，但由于缺乏适当的动物模型来模拟原发性乳腺肿瘤转移到骨的临床经验，对骨转移生物学的彻底理解仍然是难以捉摸的。 小鼠已被用作转移级联中许多单个步骤的模型，但尚未开发出整个过程的小鼠模型，即乳腺原发性肿瘤发展到其转移到远处器官如骨。 虽然，一些转基因小鼠乳腺癌模型已被证明广泛转移，但它们都没有转移到骨。neu（c-erbB 2，ERBB 2）原癌基因的过度表达和扩增与侵袭性人类乳腺癌的发展有关。 neu原癌基因特异性靶向表达至小鼠乳腺（在MMTV启动子的控制下）在205天的平均潜伏期后诱导转移性疾病。 另一方面，通常认为，在肿瘤细胞从骨髓扩展到骨之前，骨吸收是必需的。 在乳腺癌的情况下，这最后一步是由破骨细胞介导的，破骨细胞被肿瘤肽甲状旁腺激素相关肽（PTH-rp）的局部产生所刺激。 支持这一假设的一些证据是：PTH-rp阳性乳腺癌患者更容易发生骨转移。 此外，乳腺癌骨转移表达PTH-rp的病例超过90%，而非骨转移仅为17%。 还报道了PTHrp在MDA 231人乳腺癌细胞中的表达增加溶骨性骨转移，而PTHrp的中和抗体显著抑制溶骨性骨转移。 因此，PTHrp在骨转移过程中起重要作用。总之，临床和实验数据都表明，如果neu表达乳腺癌细胞被改造成过表达PTHrp，它们可能能够归巢到骨并转移。 基于这一假设，本提案的总体目标是建立乳腺癌骨转移的随机模型，并确定乳腺癌转移进展期间介导ERBB 2和PTHrp活性的下游靶点。 我们建议在体外，使用细胞培养模型和在体内通过产生neu/PTHrp表达双基因小鼠来测试这一假设。 我们的具体目标是：具体目标1：使用经工程改造过表达neu和PTHrp的体外小鼠细胞培养模型，建立乳腺癌从乳腺转移至骨的小鼠模型。 具体目标二：确定PTHrp和neu特异性靶向表达至小鼠乳腺上皮细胞是否促进这些动物的骨转移形成。开发过表达原癌基因neu并被工程化以过度产生PTHrp的这种模型将最接近地模拟人乳腺癌进展，并为治疗剂的临床前研究提供改进的模型。 它还将提供一个独特的平台，以研究乳腺癌向骨转移的位点特异性细胞事件。

Even though 70 percent patients dying with breast cancer have osteolytic bone metastasis, a thorough understanding of the biology of skeletal metastasis remains elusive due to the lack of appropriate animal models that mimic the clinical experience of metastasis of primary breast tumors to the bone. The mouse has been used as a model for a number of individual steps in the metastatic cascade, but a mouse model for the entire process i.e. development of the primary tumor in the breast to its metastases to distant organs like bone, has not been developed. Although, some transgenic mouse models of breast cancer have been shown to metastasize extensively, none of them metastasize to the bone. Overexpression and amplification of the neu (c-erbB2, ERBB2) proto-oncogene have been implicated in the development of aggressive human breast cancer. Targeted expression of the neu proto-oncogene specifically to the mouse mammary gland (under the control of a MMTV promotor) induces metastatic disease after a mean latency of 205 days. On the other hand, it is generally recognized that ostoclastic bone resorption is necessary before expansion of tumor cells from bone marrow to bone. In the case of breast cancer, this final step is mediated by osteoclasts that are stimulated by local production of the tumor peptide parathyroid hormone - related peptide (PTH-rp). Some of the evidence which supports this hypothesis are: patients with PTH-rp positive breast carcinomas are more likely to develop bone metastasis. In addition, breast carcinoma metastatic to bone express PTH-rp in greater than 90 percent of cases, compared with only 17 percent of metastases to nonbone sites. It has also been reported that expression of PTHrp in MDA 231 human breast cancer cells increased osteolytic bone metastasis while neutralising antibodies to PTHrp markedly suppressed them. Thus, PTHrp plays an important role in the process of bone metastasis. Taken together, both the clinical and experimental data suggest that if neu-expressing breast carcinoma cells are engineered to overexpress PTHrp, they may be able to home to the bone and metastasize. With this hypothesis, the overall objective of this proposal is to develop a stochastic model of breast cancer metastasis to the bone and to identify downstream targets that mediate the activity of ERBB2 and PTHrp during breast cancer metatatic progression. We propose to test this hypothesis both in vitro, using a cell culture model and in vivo by generating neu/PTHrp expressing bigenic mice. Our specific aims are: Specific Aim 1: To develop a mouse model of breast cancer metastasis from the mammary gland to the bone using an in vitro mouse cell culture model that is engineered to overexpress both neu and PTHrp. Specific Aim 2: To determine, if, targeted expression of PTHrp and neu specifically to the mouse mammary epithelial cells facilitates bone metastases formation in these animals. Development of such a model that overexpresses proto-oncogene neu and is engineered to overproduce PTHrp would most closely mimic human breast cancer progresion and provide an improved model for preclinical studies of therapeutic agents. It will also provide a unique platform to investigate cellular events involved in site-specific metastasis of breast cancer to the bone.