Department of Cardiovascular Surgery, The Second Xiangya Hospital, Central South University, Changsha, China.
Department of Thoracic Surgery, The Second Xiangya Hospital, Central South University, Changsha, China.
Thorac Cancer. 2020 Jul;11(7):1848-1860. doi: 10.1111/1759-7714.13465. Epub 2020 May 12.
Calpain 1 (CAPN1) has been found to be a promoter of cancer progression. PTPN1 as a physiological target molecule of CAPN1 plays a dephosphorylated role on multiple receptor tyrosine kinases. This study aimed to reveal the effects of CAPN1/PTPN1 on malignant phenotype and EGFR-TKI resistance of lung adenocarcinoma (LUAD) cells.
A total of 84 primary LUAD tissues and paired paracancerous normal tissues were collected. Quantitative real-time PCR (qRT-PCR) and immunohistochemical (IHC) methods were used to measure the expression of CAPN1 and PTPN1 in tissues. qRT-PCR and western blot were used to detect the expressions of CAPN1, PTPN1, c-Met and PIK3R2 in cell lines. Cell counting kit-8 (CCK-8), colony formation and transwell assay were carried out to evaluate cell erlotinib resistance, proliferation, migration and invasion. Co-IP assay was used to verify the interaction between proteins. Cycloheximide (CHX) was applied to block protein synthesis.
CAPN1, c-Met and PIK3R2 were significantly upregulated and the correlation was positive in LUAD, while PTPN1 was decreased. EGFR-sensitive mutation was related to CAPN1/PTPN1. in vitro studies showed that PTPN1 can mediate dephosphorylation of c-Met and PIK3R2 by binding with both, thereby weakening cell proliferation, metastasis and erlotinib resistance, while CAPN1 could enhance the degradation of PTPN1 protein as a cancer promoter.
CAPN1 enhances the malignant behavior and erlotinib resistance of LUAD cells via degrading PTPN1 and then activating c-Met/PIK3R2, which suggests CAPN1/PTPN1 may serve as tumor markers or potential targets for diagnosis and treatment of LUAD.
Significant findings of the study Superior CAPN1 and inferior PTPN1 were related to activation of c-Met/PIK3R2 in lung adenocarcinoma. Moreover, regulations of CAPN1 and PTPN1 induced the changes of malignant behavior and erlotinib resistance. What this study adds Our findings confirmed that CAPN1/PTPN1 play crucial roles on proliferation, metastasis and erlotinib resistance of LUAD cells as c-Met/PIK3R2 regulators, and validated the regulatory mechanism of CAPN1 on PTPN1 in tumor model for the first time.
钙蛋白酶 1（CAPN1）已被发现可促进癌症进展。作为 CAPN1 的生理靶标分子，蛋白酪氨酸磷酸酶非受体型 1（PTPN1）在多种受体酪氨酸激酶上发挥去磷酸化作用。本研究旨在揭示 CAPN1/PTPN1 对肺腺癌（LUAD）细胞恶性表型和表皮生长因子受体酪氨酸激酶抑制剂（EGFR-TKI）耐药性的影响。
收集 84 例原发性 LUAD 组织及配对癌旁正常组织，采用实时定量 PCR（qRT-PCR）和免疫组化（IHC）方法检测组织中 CAPN1 和 PTPN1 的表达。采用 qRT-PCR 和 Western blot 检测细胞系中 CAPN1、PTPN1、c-Met 和 PIK3R2 的表达。细胞计数试剂盒-8（CCK-8）、集落形成和 Transwell 实验评估细胞对厄洛替尼的耐药性、增殖、迁移和侵袭能力。采用免疫共沉淀（Co-IP）实验验证蛋白间的相互作用。应用环己酰亚胺（CHX）阻断蛋白合成。
CAPN1、c-Met 和 PIK3R2 在 LUAD 中显著上调且呈正相关，而 PTPN1 下调。EGFR 敏感突变与 CAPN1/PTPN1 相关。体外研究表明，PTPN1 通过与两者结合介导 c-Met 和 PIK3R2 的去磷酸化，从而减弱细胞增殖、转移和厄洛替尼耐药性，而 CAPN1 作为一种癌症促进剂可增强 PTPN1 蛋白的降解。
CAPN1 通过降解 PTPN1 进而激活 c-Met/PIK3R2 增强 LUAD 细胞的恶性行为和厄洛替尼耐药性，提示 CAPN1/PTPN1 可作为 LUAD 的肿瘤标志物或潜在的诊断和治疗靶点。
本研究的重要发现 LUAD 中 CAPN1 和 PTPN1 的上调和下调与 c-Met/PIK3R2 的激活有关。此外，CAPN1 和 PTPN1 的调控引起了 LUAD 细胞恶性行为和厄洛替尼耐药性的改变。 本研究的新增内容 本研究证实，CAPN1/PTPN1 作为 c-Met/PIK3R2 调节剂，在 LUAD 细胞的增殖、转移和厄洛替尼耐药性中发挥关键作用，并首次验证了 CAPN1 对肿瘤模型中 PTPN1 的调控机制。
