Role of P4HA1 in Endocrine Resistance of Breast Cancer

摘要： 他莫昔芬是最古老和使用最广泛的选择性雌激素受体调节剂。福尔维斯特是一种 选择性雌激素受体下调调节剂，阻断和损害雌激素受体。福尔维斯特是 作为三苯氧胺后疾病进展后的二线治疗引入临床。内分泌治疗是 雌激素受体阳性乳腺癌患者的一线治疗。然而，30%-50%的乳房 接受五到十年治疗的癌症患者会对内分泌治疗产生抵抗力。抵抗力 内分泌治疗和复发是乳腺癌患者死亡的主要原因。脯氨酸羟化 (P4H)，是一种将脯氨酸羟基化为4-羟基脯氨酸的胶原蛋白和其他蛋白质的-X-Pro- Gly序列作为内质网中的胶原蛋白。P4HA1，P4H1的阿尔法亚单位，负责 用于多肽结合和催化活性。在本研究中，我们发现P4HA1的表达增加是 内分泌治疗耐药乳腺癌细胞与内分泌治疗敏感乳腺癌细胞的检测比较 细胞。P4HA1的高表达与腔性乳腺癌的短期无复发生存期显著相关 病人。P4HA1蛋白失活增敏内分泌耐药瘤细胞对4-羟色胺的敏感性 羟基他莫昔芬(4OHT)或弗维斯特治疗，伴随着活性氧(ROS)的增加 内质网(ER)应激。这些初步数据表明，P4HA1是一种重要的调节因子 乳腺癌的内分泌抵抗。我们假设抑制P4HA1使管腔性乳腺癌变得敏感 细胞通过诱导ROS-ER应激途径进行内分泌治疗。研究P4HA1基因对人类免疫功能的影响 内分泌抵抗，我们将介绍P4HA1蛋白在内分泌敏感型腔性乳腺癌中的表达 细胞系。我们将检测对4OHT或福维斯特治疗的抵抗力，内质网应激水平，以及ROS水平 P4HA1过表达的细胞在4OHT或FUVESTRANT处理下。我们将在中国进行救援实验 P4HA-使用ROS抑制剂沉默内分泌耐药细胞。对于活体研究，我们将使用内分泌抵抗 腔内乳腺癌细胞系正交异性异种移植模型及耐药腔内乳腺癌患者- 衍生异种移植(PDX)模型。我们将确定P4HA1的失活是否会导致这些内分泌 耐药肿瘤对他莫昔芬或福维斯特敏感，在体内治疗，并检查是否不堪重负 ER应激和ROS水平与肿瘤的内分泌敏感性有关。总而言之，这项调查将 揭示P4HA1通过调节ROS-ER应激通路在内分泌抵抗中的新的非典型作用，以及 探索P4HA1作为内分泌难治性乳腺腔内联合治疗的新靶点 癌症患者。

Abstract: Tamoxifen is the oldest and the most widely used selective estrogen receptor modulators. Fulvestrant is a selective estrogen receptor down-regulator which blocks and damages estrogen receptors. Fulvestrant is introduced to clinical as second-line therapy post disease progression after tamoxifen. Endocrine treatment is the first-line therapy for estrogen receptor positive breast cancer patients. However, 30%-50% of the breast cancer patients with five to ten years of treatment acquire resistance to endocrine therapy. Resistance to endocrine therapy and recurrence is the leading cause of mortality in breast cancer patients. Prolyl hydroxylation (P4H), is an enzyme hydroxylates proline into 4-hydroxyproline of collagens and other proteins with the -X-Pro- Gly- sequences as collagens in the endoplasmic reticulum. P4HA1, the alpha subunit of P4H1, is responsible for both peptide binding and catalytic activity. In this study, we found that increased expression of P4HA1 was detected in endocrine therapy resistant luminal breast cancer cells compared with endocrine therapy sensitive cells. High P4HA1 expression significantly correlates with short relapse-free survival in luminal breast cancer patients. Inactivation of P4HA1 protein sensitized endocrine resistance luminal cancer cells to 4- Hydroxytamoxifen (4OHT) or fulvestrant treatment, accompanied by increased reactive oxygen species (ROS) and endoplasmic reticulum (ER) stress. These preliminary data indicate that P4HA1 is an important regulator for endocrine resistance of breast cancer. We hypothesize that inhibition of P4HA1 sensitizes luminal breast cancer cells to endocrine therapy by inducing ROS-ER stress pathway. To investigate the importance of P4HA1 for endocrine resistance, we will introduce P4HA1 protein expression in endocrine sensitive luminal breast cancer cell lines. We will examine resistance to 4OHT or fulvestrant treatment, ER stress level, as well as ROS levels in P4HA1-overexpression cells under 4OHT or fulvestrant treatment. We will perform rescue experiments in P4HA-silenced endocrine resistant cells using ROS inhibitor. For the in vivo study, we will use endocrine resistant luminal breast cancer cell lines orthotropic xenograft model, and resistance luminal breast cancer patient- derived-xenograft (PDX) models. We will determine whether inactivation of P4HA1 renders these endocrine resistant tumors sensitive to tamoxifen or fulvestrant treatment in vivo, and examine whether the overwhelmed ER stress and ROS levels are associated with endocrine sensitivity in tumors. In summary, this investigation will uncover a novel uncanonical role of P4HA1 in endocrine resistance by regulation ROS-ER stress pathway, and explore P4HA1 as a novel therapeutic target for combinatorial therapy for endocrine resistant luminal breast cancer patients.