PO.MCB04.01 · 分子与细胞生物学

PERK阳性类多倍体乳腺癌细胞的特征及其与未折叠蛋白反应特征的关联

Characterization of PERK-positive polyploid-like breast cancer cells and their association with unfolded protein response signatures

编号 7311 展板 23 时间 4/22 09:00–12:00 区域 Section 22 主讲 Huseyin Kilic, MD

分会场 Hypoxic and Proteotoxic Stress Response

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作者与单位 Authors & Affiliations

LUIS DEL POZO-YAUNER¹, Veronica Ramirez-Alcantara², Elba A. Turbat-Herrera³, Hector Chavarria Bernal¹, Maha Babker¹, Huseyin Kilic¹, Rosetta Campbell¹, Bahaaeldin Youssef¹, Ajay Singh⁴, Julio Isael Perez-Carreon⁵, Wei Yang⁶, Guillermo A. Herrera¹

¹Pathology, University of South Alabama College of Medicine, Mobile, AL,²USA Health Mitchell Cancer Institute, Mobile, AL,³University of South Alabama College of Medicine, Mobile, AL,⁴SOM-Cell and Molecular Biology, University of Mississippi Medical Center and Cancer Center and Research Institute, Jackson, MS,⁵National Institute of Genomic Medicine, Mexico City, Mexico,⁶Bruker Spatial Biology, Seattle, WA

摘要 Abstract

中文摘要

内质网应激激活未折叠蛋白反应（UPR），这是一条与乳腺癌（BC）进展、化疗耐药和多倍体巨大癌细胞（PGCC）形成相关的通路。在一项初步研究中，我们鉴定出一小群在免疫组织化学（IHC）中对磷酸化PERK抗体强阳性的乳腺癌细胞，其中一些表现出非典型有丝分裂和PGCC样形态。由于PGCC与肿瘤侵袭性和不良预后相关，我们假设强PERK阳性的BC细胞代表PGCC或其前体，并在具有活跃未折叠蛋白反应（UPR）信号的肿瘤中富集。本研究旨在明确BC中PERK阳性BC细胞的频率，评估其与UPR激活的关联，并鉴定与含有这些细胞的肿瘤相关的转录组和蛋白质组特征。使用针对磷酸化PERK、ATF6和磷酸化IRE1的抗体，通过IHC分析了131名BC患者（96例TNBC，35例非TNBC）的FFPE组织。所有样本均在治疗前采集。PERK阳性BC细胞被定义为具有强（3+）胞质和核内磷酸化PERK染色的肿瘤细胞。IHC提示PERK阳性BC细胞与TNBC表型肿瘤之间存在关联。这些细胞代表一个异质性群体，在细胞形态、大小和有丝分裂状态方面存在广泛差异。基于IHC结果，选择了21个肿瘤（13例TNBC，8例非TNBC）进行nCounter多组学分析，分别使用BC360和MO蛋白面板进行转录组和蛋白质组分析。11个肿瘤含有丰富的PERK阳性BC细胞，而10个肿瘤含有很少或没有。纳入了三份正常乳腺组织作为对照。当将含有丰富PERK阳性BC细胞的乳腺癌肿瘤与不含的进行比较时，388个基因差异表达（p值阈值0.05），其中296个p值 < 0.01。其中，SLPI、S100A7、SERPINB5、KIT和PHGDH显示表达增加最显著（log倍数变化>2.0），而132个基因，包括TFF1、PIP、TFF3、CDCA8、PAX5、IL24和VEGFD，显示表达降低最显著（log倍数变化<-2.0）。当将含丰富PERK+细胞的BC肿瘤与正常乳腺组织比较时，315个基因差异表达（p值阈值0.05），其中184个p值 < 0.01。其中，28个基因，包括SPP1、KIFC1、CDK1、TOP2A、RRM2、CENPF和MYBL2的log倍数变化>2.0，而94个基因，包括APOD、ZBTB16、PDK4、RELN、TFF3、TIMP4、EGF和VEGFD的log倍数变化<-2.0。含有丰富PERK阳性BC细胞的肿瘤显示出未折叠蛋白反应激活的明确证据，以及与侵袭性行为、代谢重编程和分化丧失相关的转录组特征。本研究由阿拉巴马州乳腺癌研究基金会（BCRFA）授予LPY的资助支持。

查看英文原文 English abstract

Endoplasmic reticulum stress activates the unfolded protein response (UPR), a pathway linked to breast cancer (BC) progression, chemoresistance, and the formation of polyploid giant cancer cells (PGCCs). In a preliminary study, we identified a small subpopulation of breast cancer cells that were strongly positive for the phospho-PERK antibody in immunohistochemistry (IHC), some of which showed atypical mitosis and PGCC-like morphology. Because PGCCs are associated with tumor aggressiveness and poor outcomes, we hypothesize that the strongly PERK-positive BC cells represent PGCCs or their precursors and are enriched in tumors with active unfolded protein response (UPR) signaling. This study aimed to define the frequency of PERK-positive BC cells in BC, assess their association with UPR activation, and identify transcriptomic and proteomic features linked to tumors containing these cells. FFPE tissues from 131 BC patients (96 TNBC, 35 non-TNBC) were analyzed by IHC using antibodies against phospho-PERK, ATF6, and phospho-IRE1. All samples were collected before treatment. PERK-positive BC cells were defined as tumor cells with strong (3+) cytoplasmic and nuclear phospho-PERK staining. The IHC indicates an association between PERK-positive BC cells and tumors with a TNBC phenotype. These cells represent an heterogeneous population with broad variation in cell morphology and size, and mitosis status. Based on IHC results, 21 tumors (13 TNBC, 8 non-TNBC) were selected for nCounter multiomics, using BC360 and MO protein panel for transcriptomic and proteomic analysis, respectively. Eleven tumors contained abundant PERK-positive BC cells, while ten had few or none. Three normal breast tissues were included as controls. When breast cancer tumors with abundant PERK-positive BC cells were compared with those without, 388 genes were differentially expressed (p-value threshold 0.05), of which 296 had p-values < 0.01. Of them, SLPI, S100A7, SERPINB5, KIT, and PHGDH showed the most significant increase in expression (log fold-change >2.0), while 132 genes, including TFF1, PIP, TFF3, CDCA8, PAX5, IL24, and VEGFD, showed the most significant decrease in expression (log fold-change <-2.0). When BC tumors with abundant PERK+ cells were compared with normal breast tissue, 315 genes were differentially expressed (p-value threshold 0.05), of which 184 had p-values < 0.01. Of them, 28, including SPP1, KIFC1, CDK1, TOP2A, RRM2, CENPF, and MYBL2 had a log fold-change >2.0, while 94 genes, including APOD, ZBTB16, PDK4, RELN, TFF3, TIMP4, EGF, and VEGFD had a log fold-change <-2.0. Tumors containing abundant PERK-positive BC cells show clear evidence of unfolded protein response activation and a transcriptomic profile associated with aggressive behavior, metabolic reprogramming, and loss of differentiation. This study was funded by a grant from the Breast Cancer Research Foundation of Alabama (BCRFA) to LPY.

利益披露 Disclosure

L. Del pozo-yauner, None.. H. Chavarria Bernal, None.. M. Babker, None.. H. Kilic, None.. R. Campbell, None.. B. Youssef, None.. A. Singh, None.. J. I. Perez-Carreon, None.. W. Yang, None.. G. A. Herrera, None.

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