Gli Control of PTH-rP and Osteolysis in Breast Cancer

描述（由申请人提供）：甲状旁腺激素相关肽（PTH-rP）不仅是恶性肿瘤（HHM）的体液性高钙血症的致病因素，也是转移性乳腺癌相关的局部骨溶解的致病因素。在这些情况下，癌细胞过度表达PTH-rP的原因尚不清楚。在乳腺癌中，骨转移部位几乎普遍表达PTH-rP。骨中PTH-rP转录的药理学抑制可减少该部位的骨溶解和肿瘤负荷（Gallwitz等，2002）。了解PTH-rP在癌细胞中过度表达的机制，将有助于确定药物的分子靶点，从而有效预防或治疗乳腺癌和其他与PTH-rP表达增加相关的癌症。我们假设Gli家族转录因子，脊椎动物细胞中的刺猬信号分子，是转移性人乳腺癌细胞中PTH-rP表达的重要调节因子，迄今尚未被研究。我们认为Gli家族成员在调节PTH-rP表达方面具有不同的功能。Gli2是一种强大的转录激活剂，而Gli3的截断形式（由特定的E3泛素连接酶β - trcp和随后的蛋白酶体加工产生）是PTH-rP转录的强烈抑制因子。（根据我们的初步数据，Gli1和Gli2没有被加工成截断的抑制因子形式，全长Gli1和Gli3对PTH-rP转录没有显著影响）。我们的初步数据表明，人乳腺癌细胞过表达Gli2， Gli2和截短的Gli3短暂转染这些细胞可调节PTH-rP的表达。乳腺癌细胞中内源性Gli2的表达与PTH-rP的表达和引起高钙血症或骨溶解的能力相关。稳定转染Gli2的癌细胞在体内导致骨溶解增强。我们计划通过确定Gli2和截短的Gli3是否增强或减少人乳腺癌细胞体内诱导的PTH-rP表达和骨溶解来验证我们的假设，确定Gli2增强PTH-rP转录的分子机制，确定Gli2和Gli3的E3连接酶β - trcp功能丧失对PTH-rP表达和骨溶解的影响，以及Gli2和截短的Gli3在导致HHM的其他过表达PTH-rP的癌细胞中的作用。因此，我们认为Gli家族转录调控因子是调节乳腺癌细胞中PTH-rP表达以及随后乳腺癌介导的骨溶解的重要分子途径。

DESCRIPTION (provided by applicant): Parathyroid hormone-related peptide (PTH-rP) is the causative factor not only in humoral hypercalcemia of malignancy (HHM), but also in the local osteolysis associated with metastatic breast cancer. The reasons for overexpression of PTH-rP by cancer cells in these situations are unknown. In breast cancer, there is almost universal expression of PTH-rP in bone metastatic sites. Pharmacologic inhibition of PTH-rP transcription in bone causes reduction in osteolysis and tumor burden in that site (Gallwitz et al., 2002). Understanding the mechanisms responsible for PTH-rP overexpression by cancer cells could thus lead to identification of molecular targets for drugs that could be effective in the prevention or treatment of breast cancer and other cancers that are associated with increased PTH-rP expression. We hypothesize that the Gli family of transcription factors, the hedgehog signaling molecules in vertebrate cells, are important heretofore uninvestigated regulators of PTH-rP expression in metastatic human breast cancer cells. We propose that Gli family members have distinct and separate functions with respect to regulating PTH-rP expression. Gli2 is a powerful transcriptional activator, and the truncated form of Gli3 (produced by the actions of the specific E3 ubiquitin ligase beta-TrCP and subsequent proteasomal processing) is a strong repressor of PTH-rP transcription. (Gli1 and Gli2 are not processed to truncated repressor forms, and full length Gli1 and Gli3 have no significant effects on PTH-rP transcription according to our Preliminary Data). Our preliminary data suggests that human breast cancer cells overexpress Gli2, and that transient transfection of these cells with Gli2 and truncated Gli3 regulate PTH-rP expression. Expression of endogenous Gli2 in breast cancer cells correlates with PTH-rP expression and capacity to cause hypercalcemia or osteolysis. Cancer cells stably transfected with Gli2 cause enhanced osteolysis in vivo. We plan to test our hypothesis by determining if Gli2 and truncated Gli3 enhance or reduce PTH-rP expression and osteolysis induced by human breast cancer cells in vivo, identify the molecular mechanisms whereby Gli2 enhances PTH-rP transcription, determine the effects of loss of function of Gli2 and the E3 ligase for Gli3, beta-TrCP, on PTH-rP expression and osteolysis, and the role of Gli2 and truncated Gli3 in other PTH-rP overexpressing cancer cells that cause HHM. We propose therefore that the Gli family of transcriptional regulators represents an important molecular pathway that regulates PTH-rP expression in breast cancer cells, and subsequent breast cancer-mediated osteolysis.