miR-130a-3p通过HGF/MET信号通路抑制乳腺癌细胞MCF-7 侵袭

目的：探究miR-130a-3p 通过HGF/MET信号通路调控上皮间质转化（epithelial-mesenchymal transition,EMT）影响乳腺癌细胞侵袭转移的分子机制。方法：收集承德医学院附属医院2018 年1 月至10 月收治的22 例乳腺癌患者癌组织和配对癌旁组织标本,乳腺癌细胞系（MCF-7、MDA-MB-231 和MDA-MB-453）和正常乳腺上皮细胞MCF10A来自承德医学院基础研究所,然后采用qPCR检测组织和细胞系中miR-130a-3p 的表达情况;将实验分为对照组、miR-130a-3p mimics 组、miR-130a-3p inhibitor组、PHA665752（MET小分子抑制剂）转染组及共转PHA665752+miR-130a-3p inhibitor 组,然后采用CCK-8 法和Transwell 实验分别检测MCF-7 细胞增殖活力、侵袭和迁移能力;WB实验检测MCF-7 细胞EMT和HGF/MET信号通路相关蛋白的表达情况;此外,采用双荧光素酶报告基因检测miR-130a-3p 与MET之间的靶向关系。结果：miR-130a-3p 在乳腺癌组织和细胞系中呈低表达;过表达miR-130a-3p 可抑制MCF-7 细胞增殖、侵袭、迁移和EMT;而抑制miR-130a-3p 出现相反的结果。双荧光素酶报告基因结果证实miR-130a-3p 靶向下调MET的表达水平,且miR-130a-3p 负调控HGF/MET信号通路的表达;进一步实验证明,miR-130a-3p 通过阻断HGF/MET信号通路抑制MCF-7 细胞增殖、侵袭、迁移和EMT。结论：miR-130a-3p 通过阻断HGF/MET信号通路抑制MCF-7 细胞EMT过程,进而抑制MCF-7 细胞侵袭转移。     

MET／HGF 抑制剂在肿瘤治疗中的研究进展

MET 是一种由 c-MET 原癌基因编码的蛋白产物,为肝细胞生长因子（hepatocyte growth factor,HGF）受体,位于细胞膜上,具有酪氨酸激酶活性,活化后最终引起细胞生物学行为改变,如细胞生长、细胞增殖、细胞运动和迁徙、细胞侵袭和促进凋亡。异常的 MET 表达在实体瘤和血液恶性肿瘤中广泛存在,具体的分子机制包括 MET 受体的过表达、基因扩增、基因突变或选择性剪接。随着对 MET 和HGF 基础研究的突破,目前已开发出 MET 抑制剂,包括：HGF 单克隆抗体、与 HGF 竞争性结合 MET 的配体、选择性和非选择性的 MET-TKI。部分安全有效的 MET 抑制剂也已进入了后期的临床研究,作者就 MET／HGF 在肿瘤中的作用及 MET 抑制剂的临床研究成果作一综述。     

非小细胞肺癌中 HGF/c-Met 信号通路与 EGFR-TKI 获得性耐药的关系

表皮生长因子受体酪氨酸激酶抑制剂（EGFR-TKI）为首的靶向药物在晚期非小细胞肺癌患者治疗中取得巨大进展。然而,获得性耐药的出现是其不可避免的结果。肝细胞生长因子（HGF）／c-Met信号通路参与多种肿瘤细胞的形成、迁徙、血管生成等重要细胞进程。该信号通路的异常激活在EGFR-TKI 获得性耐药中发挥了重要作用。实验表明,HGF ／c-Met 信号通路抑制剂可使部分 EGFR-TKI获得性耐药患者临床获益。     

PARP抑制剂在乳腺癌治疗中的研究进展

PARP抑制剂作为一种新型的小分子靶向药物,已经在乳腺癌中进行了广泛的研究,其单药在早期及晚期乳腺癌患者中已经显示出了可观的疗效,但长期单药应用所产生的PARP耐药无法避免。研究表明,PARP抑制剂与其他信号通路靶向药物,如免疫检查点抑制剂、PI3K/AKT通路抑制剂等药物的联合应用可能增强其肿瘤杀伤作用。全文就PARP抑制剂在早期及晚期乳腺癌中的单药及联合应用的研究进展进行综述。     

B细胞受体信号通路抑制剂在淋巴瘤中的应用及耐药机制研究进展:第57届美国血液学会年会报道

近年来,B细胞受体(BCR)信号通路抑制剂在复发、难治B细胞非霍奇金淋巴瘤(B-NHL)患者治疗中取得了振奋人心的疗效,大量BCR信号通路抑制剂单用或联合其他传统化疗药物、免疫调节剂、单克隆抗体及其他激酶抑制剂的临床研究正在如火如荼地开展.此外,已有研究发现了BCR信号通路抑制剂耐药的情况,其发生机制及如何克服耐药已引起关注.文章结合2015年第57届美国血液学会(ASH)年会报道的BCR信号通路抑制剂单药及联合用药的最新研究进展及其耐药机制的相关研究进行介绍,以期为临床治疗提供帮助.     

CDK4/6抑制剂联合mTOR抑制剂在乳腺癌中的作用机制

CDK-RB-E2F通路和PI3K-AKT-mTOR通路对乳腺癌耐药机制起到了关键作用。通过对两个通路的研究发现,在治疗激素受体阳性的乳腺癌时,CDK4/6抑制剂与内分泌药物联合使用可以提高患者的生存率,mTOR抑制剂也显示出抗肿瘤的实力。最近的研究表明,mTOR抑制剂和CDK4/6抑制剂联合使用可以进一步抑制CDK-RB-E2F通路激活,协同控制肿瘤细胞增殖。同时发现CDK4/6抑制剂耐药患者的CDK-RB-E2F通路重新激活,仍对mTOR抑制剂敏感。还有研究表明CDK4/6抑制剂和mTOR抑制剂可以通过自噬作用协同控制肿瘤的发生。本文针对两药联合在乳腺癌中的作用机制进行综述。     

miR-34a调控肺癌EGFR耐药细胞株中MET信号通路探讨

[目地]研究肺癌耐药细胞株HCC827/GR中MET信号通路的调控,明确肺癌获得性耐药的分子机制。[方法]使用HCC827细胞[epidermal growth factor receptor（EGFR）基因19外显子缺少的肺腺癌细胞株],在此细胞的基础上培养吉非替尼耐药细胞株。检测耐药细胞株中MET的表达,并使用RT-PCR的方法检测miR-34a的表达;使用TargetScan等生物信息学软件预测miR-34a的下游靶点,并在细胞内验证miR-34a是否可以调控MET的表达。[结果]与HCC827细胞相比,HCC827/GR细胞株对吉非替尼明显耐药。在耐药HCC827/GR细胞株中,miR-34a低表达,MET高表达,而在HCC827细胞株中,miR-34a高表达,MET低表达。Target Scan生物信息学软件提示,MET是miR-34a的下游靶点之一。将携带荧光报告基团和MET 3′UTR的载体与miR-34a共转染入细胞后荧光度下降。[结论]miR-34a可能通过调控靶基因MET而参与EGFR-TKI的获得性耐药。     

MET基因改变的非小细胞肺癌靶向治疗耐药机制及治疗策略

间质表皮转化因子（mesenchymal to epithelial transition factor, MET）基因改变参与了非小细胞肺癌的增殖、侵袭和转移。MET-酪氨酸激酶抑制剂（tyrosine kinase inhibitors, TKIs）已获批用于MET基因改变的非小细胞肺癌，而这些药物的耐药不可避免。MET-TKIs的分子耐药机制错综复杂，尚不完全清楚。本文主要对这些MET-TKIs的潜在耐药机制进行综述，以期为MET基因改变的患者提供合理的治疗思路。     

Wnt信号通路与慢性粒细胞白血病

 慢性粒细胞白血病作为血液系统的恶性增殖性肿瘤，其主要发病机制为BCR-ABL1融合基因形成，因此针对BCR-ABL1基因的靶向药物TKI为其治疗带来了希望，但TKI无法完全消除CML患者体内的LSCs，CML患者容易出现TKI药物耐药及治疗后复发。随着Wnt信号通路在实体瘤中的应用，大量研究发现Wnt信号通路同样在CML中起到重要作用，我们就Wnt信号通路对CML的影响作一综述。     

曲妥珠单抗获得性耐药机制及应对策略

摘 要：曲妥珠单抗是靶向人表皮生长因子2(HER2)的单克隆抗体药物,对HER2表达阳性乳腺癌及胃癌治疗效果确切。然而多数患者在用药1年内出现了获得性耐药,导致其疗效降低甚至无效。研究表明,信号通路异常激活;表皮生长因子受体(EGFR)及其配体异常表达及肿瘤诱发上皮—间质转化(EMT)均可成为曲妥珠单抗耐药的重要机制,联合使用PI3K/AKT/mTOR通路抑制剂及其相关生存信号通路抑制剂可逆转曲妥珠单抗耐药。本文就曲妥珠单抗耐药机制及逆转耐药策略进行综述。     

Single-agent and combination therapeutic strategies to inhibit hepatocyte growth factor/MET signaling in cancer

Receptor tyrosine kinases are often aberrantly activated in human malignancies and contribute to cancer development and progression. Specific receptor tyrosine kinase inhibitors have been shown to be clinically effective therapies in subsets of cancer patients with either hematologic or solid tumors. Activation of the hepatocyte growth factor (HGF)/MET signaling pathway has been found to play a critical role in oncogenesis, cancer metastasis, and drug resistance. These observations have led to the development of agents that can effectively inhibit HGF/MET signaling through direct inhibition of the receptor (anti-MET antibodies), through inactivation of its ligand HGF (AMG102, L2G7), by interfering with HGF binding to MET (NK4), or by inhibiting MET kinase activity (PHA-665752 and SU11274). Moreover, the combination of anti-MET therapeutic agents with either signal transduction inhibitors (ERBB family or mTOR inhibitors) or with cytotoxic chemotherapy has been evaluated in preclinical models. These studies provide insight into the rational development of combination therapeutic strategies that can be evaluated in clinical trials. This review will discuss different strategies of MET inhibition with a specific focus on combination therapeutic approaches.     

Hepatocyte growth factor/MET in cancer progression and biomarker discovery

Signaling driven by hepatocyte growth factor (HGF) and MET receptor facilitates conspicuous biological responses such as epithelial cell migration, 3‐D morphogenesis, and survival. The dynamic migration and promotion of cell survival induced by MET activation are bases for invasion–metastasis and resistance, respectively, against targeted drugs in cancers. Recent studies indicated that MET in tumor‐derived exosomes facilitates metastatic niche formation and metastasis in malignant melanoma. In lung cancer, gene amplification‐induced MET activation and ligand‐dependent MET activation in an autocrine/paracrine manner are causes for resistance to epidermal growth factor receptor tyrosine kinase inhibitors and anaplastic lymphoma kinase inhibitors. Hepatocyte growth factor secreted in the tumor microenvironment contributes to the innate and acquired resistance to RAF inhibitors. Changes in serum/plasma HGF, soluble MET (sMET), and phospho‐MET have been confirmed to be associated with disease progression, metastasis, therapy response, and survival. Higher serum/plasma HGF levels are associated with therapy resistance and/or metastasis, while lower HGF levels are associated with progression‐free survival and overall survival after treatment with targeted drugs in lung cancer, gastric cancer, colon cancer, and malignant melanoma. Urinary sMET levels in patients with bladder cancer are higher than those in patients without bladder cancer and associated with disease progression. Some of the multi‐kinase inhibitors that target MET have received regulatory approval, whereas none of the selective HGF‐MET inhibitors have shown efficacy in phase III clinical trials. Validation of the HGF‐MET pathway as a critical driver in cancer development/progression and utilization of appropriate biomarkers are key to development and approval of HGF‐MET inhibitors for clinical use.     

The Role of MET Receptor Tyrosine Kinase in Non‐Small Cell Lung Cancer and Clinical Development of Targeted Anti‐MET Agents

A better understanding of the pathophysiology and evolution of non‐small cell lung cancer (NSCLC) has identified a number of molecular targets and spurred development of novel targeted therapeutic agents. The MET receptor tyrosine kinase and its ligand hepatocyte growth factor (HGF) are implicated in tumor cell proliferation, migration, invasion, and angiogenesis in a broad spectrum of human cancers, including NSCLC. Amplification of MET has been reported in approximately 5%–22% of lung tumors with acquired resistance to small‐molecule inhibitors of the epidermal growth factor receptor (EGFR). Resistance to EGFR inhibitors is likely mediated through downstream activation of the phosphoinositide 3‐kinase /AKT pathway. Simultaneous treatment of resistant tumors with a MET inhibitor plus an EGFR inhibitor can abrogate activation of downstream effectors of cell growth, proliferation, and survival, thereby overcoming acquired resistance to EGFR inhibitors. Development and preclinical testing of multiple agents targeting the HGF–MET pathway, including monoclonal antibodies targeting HGF or the MET receptor and small‐molecule inhibitors of the MET tyrosine kinase, have confirmed the crucial role of this pathway in NSCLC. Several agents are now in phase III clinical development for the treatment of NSCLC. This review summarizes the role of MET in the pathophysiology of NSCLC and in acquired resistance to EGFR inhibitors and provides an update on progress in the clinical development of inhibitors of MET for treatment of NSCLC.     

Impaired ligand‐dependent MET activation caused by an extracellular SEMA domain missense mutation in lung cancer

Aberrant activation of the MET/hepatocyte growth factor (HGF) receptor participates in the malignant behavior of cancer cells, such as invasion‐metastasis and resistance to molecular targeted drugs. Many mutations in the MET extracellular region have been reported, but their significance is largely unknown. Here, we report the dysregulation of mutant MET originally found in a lung cancer patient with Val370 to Asp370 (V370D) replacement located in the extracellular SEMA domain. MET‐knockout cells were prepared and reconstituted with WT‐MET or V370D‐MET. HGF stimulation induced MET dimerization and biological responses in cells reconstituted with WT‐MET, but HGF did not induce MET dimerization and failed to induce biological responses in V370D‐MET cells. The V370D mutation abrogated HGF‐dependent drug resistance of lung cancer cells to epidermal growth factor receptor‐tyrosine kinase inhibitors (EGFR‐TKI). Compared with WT‐MET cells, V370D‐MET cells showed different activation patterns in receptor tyrosine kinases upon exposure to survival/growth‐stressed conditions. Surface plasmon resonance analysis indicated that affinity between the extracellular region of V370D‐MET and HGF was reduced compared with that for WT‐MET. Further analysis of the association between V370D‐MET and the separate domains of HGF indicated that the SP domain of HGF was unchanged, but its association with the NK4 domain of HGF was mostly lost in V370D‐MET. These results indicate that the V370D mutation in the MET receptor impairs the functional association with HGF and is therefore a loss‐of‐function mutation. This mutation may change the dependence of cancer cell growth/survival on signaling molecules, which may promote cancer cell characteristics under certain conditions.     

Seminars in clinical pharmacology: an introduction to MET inhibitors for the medical oncologist

MET is a tyrosine kinase receptor for hepatocyte growth factor (HGF), primarily expressed on epithelial cells; the activation of MET induces several biological responses relevant for the development and growth of many human cancers. Several human malignancies present altered expression of MET and this is usually associated with poor prognosis and aggressive phenotype. The majority of MET inhibitors in clinical development target directly the receptor through the use of monoclonal antibodies (MAbs) or through small molecule inhibitors of MET kinase activity; small molecule inhibitors are very potent but less specific than MAbs. MET inhibitors are of great clinical interest because of the extensive crosstalk of the HGF/MET axis with many other signaling pathways, including growth factor-dependent pathways (like PI3K/AKT/mTOR,RAS/RAF/ERK) and vascular endothelial growth factor (VEGF) axis. In preclinical studies, the treatment with MET inhibitors could prevent or reverse resistance to inhibitors of growth factor-dependent signaling; this hypothesis is currently tested in phase III trials with anti-epidermal growth factor receptor (EGFR) inhibitors in non-small-cell lung cancer (NSCLC). Based on preclinical and preliminary clinical results, a rational strategy for the clinical development of MET antagonists should include a selection of the tumors with MET overexpression, the identification of prognostic/predictive biomarkers, the evaluation of combinations with anti-VEGF compounds.     

The emerging role of MET/HGF inhibitors in oncology

The N-methyl-N′-nitroso-guanidine human osteosarcoma transforming gene (MET) receptor tyrosine kinase and its ligand hepatocyte growth factor (HGF) control cellular signaling cascades that direct cell growth, proliferation, survival, and motility. Aberrant MET/HGF activation has been observed in many tumor types, can occur by multiple mechanisms, and promotes cellular proliferation and metastasis via growth factor receptors and other oncogenic receptor pathways. Thus, MET/HGF inhibition has emerged as targeted anticancer therapies. Preclinically, neoplastic and metastatic phenotypes of several tumor cells, including non-small cell lung cancer, hepatocellular carcinoma, and gastric cancer, were abrogated by MET inhibition. Ongoing clinical development with tivantinib, cabozantinib, onartuzumab, crizotinib, rilotumumab, and ficlatuzumab has shown encouraging results. These trials have established a key role for MET in a variety of tumor types. Evidence is emerging for identification of aberrant MET activity biomarkers and selection of patient subpopulations that may benefit from targeted MET and HGF inhibitor treatment.     

Multiple mutations and bypass mechanisms can contribute to development of acquired resistance to MET inhibitors

Therapies targeting receptor tyrosine kinases have shown efficacy in molecularly defined subsets of cancers. Unfortunately, cancers invariably develop resistance, and overcoming or preventing resistance will ultimately be key to unleashing their full therapeutic potential. In this study, we examined how cancers become resistant to MET inhibitors, a class of drugs currently under clinical development. We utilized the highly sensitive gastric carcinoma cell line, SNU638, and two related MET inhibitors PHA-665752 and PF-2341066. To our surprise, we observed at least two mechanisms of resistance that arose simultaneously. Both resulted in maintenance of downstream PI3K (phosphoinositide 3-kinase)-AKT and MEK (MAP/ERK kinase)-ERK signaling in the presence of inhibitor. One mechanism, observed by modeling resistance both in vitro and in vivo, involved the acquisition of a mutation in the MET activation loop (Y1230). Structural analysis indicates that this mutation destabilizes the autoinhibitory conformation of MET and abrogates an important aromatic stacking interaction with the inhibitor. The other cause of resistance was activation of the epidermal growth factor receptor (EGFR) pathway due to increased expression of transforming growth factor α. Activation of EGFR bypassed the need for MET signaling to activate downstream signaling in these cells. This resistance could be overcome by combined EGFR and MET inhibition. Thus, therapeutic strategies that combine MET inhibitors capable of inhibiting Y1230 mutant MET in combination with anti-EGFR-based therapies may enhance clinical benefit for patients with MET-addicted cancers. Importantly, these results also underscore the notion that a single cancer can simultaneously develop resistance induced by several mechanisms and highlight the daunting challenges associated with preventing or overcoming resistance.     

The promise of selective MET inhibitors in non-small cell lung cancer with MET exon 14 skipping

Dysregulated activation of the MET tyrosine kinase receptor is implicated in the development of solid tumors and can arise through several mechanisms, including gene amplification, overexpression of the receptor and/or its ligand hepatocyte growth factor (HGF), and the acquisition of activating mutations. The most common activating mutations cause exon 14 to be skipped during MET mRNA splicing. This in-frame deletion, known as MET exon 14, results in production of a shortened receptor that lacks a juxtamembrane domain but retains affinity for HGF. However, the negative regulatory function located within this protein sequence is lost, leading to receptor accumulation on the cell surface and prolonged activation by HGF. MET mutations causing exon 14 skipping appear to be true oncogenic drivers and occur in patients and tumors with distinct characteristics.Increasing evidence suggests that tumors carrying such mutations are sensitive to MET inhibition, raising the hope that selective MET inhibitors will provide patients with optimal anticancer activity with minimal toxicity.We discuss the prospects for selective MET inhibitors in the treatment of non-small cell lung cancer harboring MET exon 14 skipping.     

MET signaling: novel targeted inhibition and its clinical development in lung cancer

MET is a versatile receptor tyrosine kinase within the human kinome which is activated by its specific natural ligand hepatocyte growth factor (HGF). MET signaling plays an important physiologic role in embryogenesis and early development, whereas its deregulation from an otherwise quiescent signaling state in mature adult tissues can lead to upregulated cell proliferation, survival, scattering, motility and migration, angiogenesis, invasion, and metastasis in tumorigenesis and tumor progression. Studies have shown that MET pathway is activated in many solid and hematological malignancies, including lung cancer, and can be altered through ligand or receptor overexpression, genomic amplification, MET mutations, and alternative splicing. The MET signaling pathway is known to be an important novel target for therapeutic intervention in human cancer. A number of novel therapeutic agents that target the MET/HGF pathway have been tested in early-phase clinical studies with promising results. Phase 3 studies of MET targeting agents have just been initiated. We will review the MET signaling pathway and biology in lung cancer and the recent clinical development and advances of MET/HGF targeting agents with emphasis on discussion of issues and strategies needed to optimize the personalized therapy and further clinical development.     

Targeting MET in cancer: rationale and progress

Uncontrolled cell survival, growth, angiogenesis and metastasis are essential hallmarks of cancer. Genetic and biochemical data have demonstrated that the growth and motility factor hepatocyte growth factor/scatter factor (HGF/SF) and its receptor, the tyrosine kinase MET, have a causal role in all of these processes, thus providing a strong rationale for targeting these molecules in cancer. Parallel progress in understanding the structure and function of HGF/SF, MET and associated signalling components has led to the successful development of blocking antibodies and a large number of small-molecule MET kinase inhibitors. In this Review, we discuss these advances, as well as results from recent clinical studies that demonstrate that inhibiting MET signalling in several types of solid human tumours has major therapeutic value.