Ewing's Sarcoma Resistance to Immunity and Radiation

描述（由申请人提供）：尤文氏肉瘤是一种非常痛苦的骨癌（更罕见的是软组织），袭击儿童和青少年。尽管接受了手术、化疗和放疗，但许多尤文肉瘤患者无法存活，特别是如果癌症已经转移。复发性尤文氏肉瘤往往表现出对化疗和放疗的抵抗力，这促使新疗法的发展，包括免疫疗法。然而，一些尤文氏肉瘤细胞可以逃避免疫系统的消除，这种癌症逃避的机制知之甚少，尽管有报道称主要组织相容性复合体（MHC）I类分子的表达下调，以及MHC I类表达降低与生存率低下的相关性。由于MHC I类分子的功能是将抗原（包括癌症相关抗原）呈递给细胞毒性T淋巴细胞，因此其表面表达的减少可能导致免疫逃避。我们试图描述与尤因肉瘤细胞相关的避免免疫识别的细胞变化，研究MHC分子和淀粉样前体样蛋白2（APLP 2）在这一过程中的作用。APLP 2是一个广泛表达的蛋白质家族成员，该蛋白质家族还包括淀粉样前体蛋白（APP）和神经系统限制性淀粉样前体蛋白样蛋白1（APLP 1）。APLP 2在细胞表面以跨膜形式存在，其被β-分泌酶切割导致产生约12 kDa的C-末端片段。我们的新数据表明，尤文氏肉瘤细胞系中APLP 2的表达升高降低了MHC I类分子的表面表达。放射治疗后，APLP 2表达相对较低的尤因肉瘤细胞在细胞表面的MHC I类表达增加，但APLP 2高的尤因肉瘤亚群没有。在我们的初步实验中，我们还证明了尤文氏肉瘤细胞照射后APLP 2减少了G2/M期的细胞周期停滞，并降低了具有亚G1 DNA含量的细胞比例（表明细胞凋亡）。根据我们的初步数据，我们的中心假设是APLP 2促进了 尤文肉瘤细胞。为了验证我们的中心假设，本项目的具体目标是（1）确定APLP 2在尤文肉瘤细胞抗辐射前后细胞毒性中的作用，以及（2）确定APLP 2对尤文肉瘤细胞抗辐射的贡献。总体而言，本申请中提出的研究有望为调节尤文肉瘤细胞存活的分子途径提供新的视角，涉及免疫和放疗的逃避。

DESCRIPTION (provided by applicant): Ewing's sarcoma is a very painful cancer of the bone (and, more rarely, soft tissue) that strikes children and adolescents. Despite treatment with surgery, chemotherapy, and radiation, many patients with Ewing's sarcoma do not survive, particularly if the cancer has metastasized. Recurrent Ewing's sarcoma tends to exhibit resistance to chemo- and radiotherapies, motivating the development of novel therapies, including immunotherapies. However, some Ewing's sarcoma cells can escape elimination by the immune system and the mechanisms underlying evasion by this cancer are poorly understood, although there have been reports of down-regulated expression of major histocompatibility complex (MHC) class I molecules, and correlation of decreased MHC class I expression and poor survival. Since the function of MHC class I molecules is to present antigens (including cancer-associated antigens) to cytotoxic T lymphocytes, reduction of their surface expression could cause immune evasion. We have sought to characterize cellular changes associated with Ewing's sarcoma cells that avoid immune recognition, investigating the roles of MHC molecules and amyloid precursor-like protein 2 (APLP2) in this process. APLP2 is a ubiquitously expressed member of a protein family that also includes amyloid precursor protein (APP) and the nervous system-restricted amyloid precursor-like protein 1 (APLP1). APLP2 is found in transmembrane form at the cell surface, and its cleavage by beta-secretases leads to the production of ~12 kDa C-terminal fragments. Our new data suggests that elevated expression of APLP2 in Ewing's sarcoma cell lines reduces surface expression of MHC class I molecules. Following treatment with radiation, the Ewing's sarcoma cells that had relatively low APLP2 expression had increased MHC class I expression at the cell surface, but APLP2- high Ewing's sarcoma sub-populations did not. In our preliminary experiments, we have also demonstrated that following irradiation of Ewing's sarcoma cells APLP2 reduces cell cycle arrest at G2/M and lowers the proportion of cells with sub-G1 DNA content (indicating apoptosis). On the basis of our preliminary data, our central hypothesis is that APLP2 facilitates the survival of Ewing's sarcoma cells. To test our central hypothesis, the Specific Aims of this proposed project are (1) to determine the role of APLP2 in resistance of Ewing's sarcoma cells, pre- and post-irradiation, to cytotoxicity, and (2) to determine the contribution of APLP2 to radiation resistanc by Ewing's sarcoma cells. Overall, the studies proposed in this application are expected to provide a new perspective on molecular pathways regulating Ewing's sarcoma cell survival, involving evasion of both immunity and radiotherapy.