MiR-152/PKM2/SLC7A5 axis in breast cancer development, chemo- and radiation-treatment response

乳腺癌（BCa）是世界范围内女性癌症死亡的主要原因，三阴性乳腺癌（BCa）是女性癌症死亡的主要原因。 TNBC癌细胞占BCa的15-20%。然而，较高的治疗抗性和较低的存活率 的TNBC患者仍然是BCa治疗的主要障碍。多柔比星（Dox）和放射治疗通常 用于TNBC治疗。在我们的初步研究中，我们发现TNBC的表达要低得多， miR-152的水平。Zeste同源物2（EZH 2）和DNA超甲基化的表达水平较高， 参与TNBC细胞中的miR-152抑制。了解miR-152抑制的机制， miR-152在调节癌症发展以及Dox和辐射诱导的癌症中的作用和机制 我们进行了大量的初步实验，以确定哪些分子是潜在的 miR-152直接靶向以及在Dox抗性TNBC或PDX肿瘤组织中上调。我们 发现丙酮酸激酶肌肉2（PKM 2）、溶质载体家族7成员5（SLC 7A 5）和 作为miR-152直接靶点的多聚嘧啶片段结合蛋白（PTBP 1），具有潜在的匡威能力 Dox介导的抗性。当miR-152表达增加时，PKM 2、SLC 7A 5和PTBP 1水平增加。 在TNBC细胞和PDX模型中被抑制。我们发现，miR-152抑制在 在介导TNBC代谢重编程、瓦尔堡效应开关、肿瘤生长和Dox抗性中的作用 通过PKM 2/SLC 7A 5/PTBP 1。我们发现，miR-152的过表达或PKM 2的敲低使 TNBC细胞对放射治疗更敏感。这些新发现为研究创造了新的机会 TNBC治疗抗性的机制。我们假设对化疗的抵抗和 放射治疗、代谢重编程和TNBC发展由miR-152诱导 PKM 2、SLC 7A 5和PTBP 1的抑制和上调。为了验证这一假设，我们将 通过三个目标进行实验。目的1：探讨miR-152在肿瘤细胞中的作用及机制 TNBC细胞在阿霉素和辐射诱导的抗性中的抑制;以及 通过miR-152抑制经由选择性剪接的PKM 2转换和诱导以诱导瓦尔堡效应， 治疗抗性和肿瘤生长。在目标2中，我们将确定miR- 152/SLC 7A 5/PTBP 1通路在调节代谢重编程和瓦尔堡效应开关中的作用，以及在 介导阿霉素和放射诱导的治疗抗性。在目标3中，我们将确定 miR-152抑制和PKM 2诱导通过CXCL 8表达调节肿瘤血管生成 人源化嵌合肿瘤模型;以及miR-152、PKM 2、HIF-1α、hnRNPA 3、SLC 7A 5和 PTBP 1相互关联，并与治疗反应和癌症分期相关 和生存这个R 01项目将确定治疗耐药性的机制，以及代谢 重新编程;并为未来开发TNBC的新治疗方案提供信息。

Breast cancer (BCa) is the leading cause of women cancer mortality worldwide, and triple negative breast cancer (TNBC) cells accounts for 15-20% BCa. However, higher therapeutic resistance and lower survival of TNBC patients remain the major hinder of BCa treatment. Doxorubicin (Dox) and radiation are commonly used for TNBC treatment. In our preliminary study, we found that TNBC showed much lower expression levels of miR-152. Higher expression levels of zeste homologue 2 (EZH2) and DNA hypermethylation were involved in miR-152 suppression in TNBC cells. To understand mechanism of miR-152 suppression, and role and mechanism of miR-152 in regulating cancer development and Dox- and radiation-induced resistance, we performed a lot of preliminary experiments to identify what molecules that were potential miR-152 direct targets as well as that were upregulated in Dox-resistant TNBC or PDX tumor tissues. We found that pyruvate kinase muscle 2 (PKM2), solute Carrier Family 7 Member 5 (SLC7A5), and polypyrimidine tract-binding protein (PTBP1) as direct targets of miR-152 with potential ability to converse Dox-mediated resistance. PKM2, SLC7A5 and PTBP1 levels were increased when miR-152 expression was suppressed in TNBC cells and PDX model. We found that miR-152 suppression played an important role in mediating TNBC metabolic reprogramming, Warburg effect switch, tumor growth and Dox resistance through PKM2/SLC7A5/PTBP1. We showed that overexpression of miR-152 or PKM2 knockdown rendered TNBC cells more sensitive to radiation treatment. These new findings create new opportunity to investigate mechanism of TNBC therapeutic resistance. We hypothesize that resistance to chemotherapy and radiation treatment, metabolic reprogramming, and TNBC development are induced by miR-152 suppression and upregulation of PKM2, SLC7A5, and PTBP1. To test this central hypothesis, we will perform experiments through three aims. In Aim 1, we will investigate role and mechanism of miR-152 suppression in TNBC cells in doxorubicin- and radiation-induced resistance; and role and mechanism of PKM2 switch and induction via alternative splicing by miR-152 suppression to induce Warburg effect, therapeutic resistance, and tumor growth. In Aim 2, we will determine role and mechanism of miR- 152/SLC7A5/PTBP1 pathway in regulating metabolic reprogramming and Warburg effect switch, and in mediating doxorubicin- and radiation-induced therapeutic resistance. In Aim 3, we will determine whether miR-152 suppression and PKM2 induction regulate tumor angiogenesis through CXCL8 expression using a humanized chimeric tumor model; and whether levels of miR-152, PKM2, HIF-1α, hnRNPA3, SLC7A5, and PTBP1 are correlated each other, and are correlated with therapeutic responses and with the cancer stages and survival. This R01 project will identify mechanisms of therapeutic resistance, and metabolic reprogramming; and provide information for developing new treatment option of TNBC in the future.