Molecular biology of epidermal growth factor receptor inhibition for cancer therapy

Understanding the role of the epidermal growth factor receptor (EGFR) in cellular signalling processes underlying malignancy has enabled the development of rationally designed EGFR-targeted therapeutics. Strategies have been devised to interfere with the EGFR signalling at three different levels: at the extracellular level, competing with ligand binding; at the intracellular level, inhibiting the activation of the tyrosine kinase; or at the mRNA level, modulating the expression of the EGFR protein. Each of these strategies has proven to have an antitumour effect mediated by events such as inhibition of cell proliferation, induction of apoptosis, decrease of cellular invasion and migration; and/or inhibition of angiogenesis. Furthermore, the combination of these strategies with traditional chemotherapy or radiotherapy has generally resulted in enhanced antitumour effects. Likewise, the benefit of interfering simultaneously with different signalling pathways has been documented to improve tumour growth inhibition. These preclinical results have encouraged clinical studies that led to the FDA approval of three drugs. However, finding the perfect strategy for each individual patient appears to be a limiting factor, demanding further research to be able to generate relevant molecular expression profiles on a case-to-case basis. Taken together, a successful EGFR inhibition will require a better understanding of signalling pathways in combination with the development of rationally designed effective molecules.     

Molecular biology of epidermal growth factor receptor inhibition for cancer therapy

Understanding the role of the epidermal growth factor receptor (EGFR) in cellular signalling processes underlying malignancy has enabled the development of rationally designed EGFR-targeted therapeutics. Strategies have been devised to interfere with the EGFR signalling at three different levels: at the extracellular level, competing with ligand binding; at the intracellular level, inhibiting the activation of the tyrosine kinase; or at the mRNA level, modulating the expression of the EGFR protein. Each of these strategies has proven to have an antitumour effect mediated by events such as inhibition of cell proliferation, induction of apoptosis, decrease of cellular invasion and migration; and/or inhibition of angiogenesis. Furthermore, the combination of these strategies with traditional chemotherapy or radiotherapy has generally resulted in enhanced antitumour effects. Likewise, the benefit of interfering simultaneously with different signalling pathways has been documented to improve tumour growth inhibition. These preclinical results have encouraged clinical studies that led to the FDA approval of three drugs. However, finding the perfect strategy for each individual patient appears to be a limiting factor, demanding further research to be able to generate relevant molecular expression profiles on a case-to-case basis. Taken together, a successful EGFR inhibition will require a better understanding of signalling pathways in combination with the development of rationally designed effective molecules.     

血小板源性生长因子及其受体与胃癌的靶向治疗

血小板源性生长因子(PDGF)及其受体与肿瘤的发生、新生血管的生成、控制肿瘤组织间隙压以及细胞凋亡机制等密切相关。新近研究发现,通过靶向抑制PDGF及其受体能够抑制胃癌的新血管生成和增强细胞毒性化疗药物的疗效。现综述PDGF及其受体在胃癌分子靶向治疗方面的研究进展。     

The epidermal growth factor receptor and its inhibition in cancer therapy.

Much effort has been expended in the search for inhibitors of signalling molecules that may prove to be important therapeutically in cancer. The epidermal growth factor receptor (EGFR) family and their associated ligands has been one such area extensively investigated. The complex nature of EGFR biology allows for potential opportunities for EGFR inhibitors in a number of areas of cancer therapy, including proliferative, angiogenic, invasive, and metastatic aspects. Much positive evidence of likely benefit has already been gathered from a multiplicity of laboratory experiments. Clinical trials are now urgently required to further evaluate the advantages of such agents.     

表皮生长因子受体活化与胰腺癌细胞解离关系的实验研究

目的 明确表皮生长因子受体(EGFR)活化参与胰腺癌细胞解离调节的分子机制.方法 通过免疫荧光法检测仓鼠高转移株(PC-1.0)和低转移株(PC-1)胰腺癌细胞中EGFR、活化(磷酸化)EGFR (p-EGFR)、活化(磷酸化)丝裂原活化蛋白激酶激酶2 (p-MEK1/2)及活化(磷酸化)细胞外信号调节激酶1/2 (p-ERK1/2)的表达变化及其与胰腺癌细胞解离状态变化的关系.结果 胰腺癌细胞解离因子(DF)明显诱导低转移株胰腺癌细胞(PC-1)中EGFR、p-EGFR、p-MEK1/2和p-ERK1/2的表达,同时诱导其细胞克隆解离.相反,AG1478(EGFR活化抑制剂)明显抑制高转移株胰腺癌细胞(PC-1.0)中EGFR、p-EGFR、p-MEK1/2和p-ERK1/2的表达,同时诱导PC-1.0细胞聚集成细胞克隆.结论 表皮生长因子受体活化后激活MEK/ERK信号通路,从而参与胰腺癌细胞解离的调节.     

Antitumor mechanisms of combined gastrin-releasing peptide receptor and epidermal growth factor receptor targeting in head and neck cancer

Head and neck squamous cell carcinoma (HNSCC) is characterized by epidermal growth factor receptor (EGFR) overexpression, where EGFR levels correlate with survival. To date, EGFR targeting has shown limited antitumor effects in head and neck cancer when administrated as monotherapy. We previously identified a gastrin-releasing peptide/gastrin-releasing peptide receptor (GRP/GRPR) aurocrine regulatory pathway in HNSCC, where GRP stimulates Src-dependent cleavage of EGFR proligands with subsequent EGFR phosphorylation and mitogen-activated protein kinase (MAPK) activation. To determine whether GRPR targeting can enhance the antitumor efficacy of EGFR inhibition, we investigated the effects of a GRPR antagonist (PD176252) in conjunction with an EGFR tyrosine kinase inhibitor (erlotinib). Combined blockade of GRPR and EGFR pathways significantly inhibited HNSCC, but not immortalized mucosal epithelial cell, proliferation, invasion, and colony formation. In addition, the percentage of apoptotic cells increased upon combined inhibition. The enhanced antitumor efficacy was accompanied by increased expression of cleaved poly(ADP-ribose) polymerase (PARP) and decreased phospho-EGFR, phospho-MAPK, and proliferating cell nuclear antigen (PCNA). Using reverse-phase protein microarray (RPPA), we further detected decreased expression of phospho-c-Jun, phospho-p70S6K, and phospho-p38 with combined targeting. Cumulatively, these results suggest that GRPR targeting can enhance the antitumor effects of EGFR inhibitors in head and neck cancer.     

Overexpression of the epidermal growth factor receptor in human pancreatic cancer is associated with concomitant increases in the levels of epidermal growth factor and transforming growth factor alpha.

The epidermal growth factor (EGF) receptor is activated by both EGF and transforming growth factor-alpha (TGF-alpha). Using immunohistochemical and immunoblotting techniques we now report that the EGF receptor, EGF, and TGF-alpha are found in both pancreatic acini and ducts in the normal human pancreas, and that all three proteins are expressed at higher levels in human pancreatic cancer tissues. Using in situ hybridization techniques, we also report that the mRNA encoding the EGF receptor, EGF, and TGF-alpha colocalize with their respective proteins. Northern blot analysis of total RNA indicates that, by comparison with the normal pancreas, the pancreatic tumors exhibit a 3-, 15-, and 10-fold increase in the mRNA levels encoding the EGF receptor, EGF, and TGF-alpha, respectively. Furthermore, by in situ hybridization, there is a marked increase in these mRNA moieties within the tumor mass. These findings suggest that EGF and TGF-alpha may participate in the regulation of normal pancreatic exocrine function, and that overexpression of the EGF receptor and its two principal ligands may contribute to the pathophysiological processes that occur in human pancreatic cancer.     

胃癌组织中表皮生长因子受体和环氧合酶-2的表达及其临床意义

目的 探讨表皮生长因子受体(epithelial growth factor receptor,EGFR)和环氧合酶-2(cyclooxygenase-2,COX-2)在胃癌组织中的表达及其临床意义与相互关系.方法 采用免疫组化Envision二步法检测40例胃癌组织中EGFR和COX-2的表达.结果 IEGFR表达阳性率为72.50%(29/40),COX-2阳性表达率为52.50%(21/40),二者的表达与胃癌淋巴结转移明显相关(P<0.05);②EGFR和COX-2在胃癌中的阳性表达与临床分期有关,在TNM分期中Ⅲ+Ⅳ期阳性表达率高于Ⅰ+Ⅱ期(P<0.05);③胃癌组织中COX-2表达与EGFR表达之间存在相关性(P<0.05).结论 COX-2和EGFR过度表达参与了胃癌的发生发展过程,并与其淋巴结转移和TNM分期有关;胃癌中COX-2和EGFR的表达具有正向协同性.     

Dual mitogen-activated protein kinase and epidermal growth factor receptor inhibition in biliary and pancreatic cancer

This study aimed to develop rational combinations of targeted agents against biliary and pancreatic cancers. To this end, we compared the global gene expression profile of biliary cancer cell lines with different degrees of sensibility to the epidermal growth factor receptor tyrosine kinase inhibitors gefitinib and erlotinib using the Affymetrix U133A microarray platform. A set of 32 genes, including genes involved in signal transduction pathways, cell cycle regulation, and angiogenesis, was highly overexpressed in resistant cells. Five of these genes encoded proteins in the Ras/Raf/mitogen-activated protein kinase (MAPK) pathway, a finding that was confirmed by Western blot and immunohistochemistry. Gefitinib failed to inhibit the MAPK pathway in resistant cell lines. Based on these data, we explored the activity of dual treatment with gefitinib in combination with CI-1040, a MAPK inhibitor. This strategy effectively resulted in inhibition of the MAPK signaling pathway and exerted antitumor effects in vitro and in vivo in tumors resistant to each of the agents alone. To further confirm these results, we tested the combined treatment in four tumor xenografts generated from patients with resected pancreatic cancer. Combined treatment was more effective than either single agent alone in this model. This study illustrates the value of global analysis of gene expression to rationally design combinations of mechanistic-based drugs. In addition, the data support the efficacy of combined epidermal growth factor receptor and MAPK inhibitors in biliary and pancreatic cancers, providing the basis to test this combination in the clinic.     

In vitro and in vivo targeting of hollow gold nanoshells directed at epidermal growth factor receptor for photothermal ablation therapy

Laser-induced phototherapy is a new therapeutic use of electromagnetic radiation for cancer treatment. The use of targeted plasmonic gold nanoparticles can reduce the laser energy necessary for selective tumor cell destruction. However, the ability for targeted delivery of the currently used gold nanoparticles to tumor cells is limited. Here, we describe a new class of molecular specific photothermal coupling agents based on hollow gold nanoshells (HAuNS; average diameter, approximately 30 nm) covalently attached to monoclonal antibody directed at epidermal growth factor receptor (EGFR). The resulting anti-EGFR-HAuNS exhibited excellent colloidal stability and efficient photothermal effect in the near-infrared region. EGFR-mediated selective uptake of anti-EGFR-HAuNS in EGFR-positive A431 tumor cells but not IgG-HAuNS control was shown in vitro by imaging scattered light from the nanoshells. Irradiation of A431 cells treated with anti-EGFR-HAuNS with near-infrared laser resulted in selective destruction of these cells. In contrast, cells treated with anti-EGFR-HAuNS alone, laser alone, or IgG-HAuNS plus laser did not show observable effect on cell viability. Using 111In-labeled HAuNS, we showed that anti-EGFR-HAuNS could be delivered to EGFR-positive tumors at 6.8% ID/g, and the microscopic image of excised tumor with scattering signal from nanoshells confirmed preferential delivery to A431 tumor of anti-EGFR-HAuNS compared with IgG-HAuNS. The absence of silica core, the relatively small particle size and high tumor uptake, and the absence of cytotoxic surfactant required to stabilize other gold nanoparticles suggest that immuno-HAuNS have the potential to extend to in vivo molecular therapy.     

Targeted therapy of orthotopic human lung cancer by combined vascular endothelial growth factor and epidermal growth factor receptor signaling blockade

The outcome for patients with lung cancer has not changed significantly for more than two decades. Several studies show that the overexpression of vascular endothelial growth factor (VEGF)/vascular permeability factor and epidermal growth factor (EGF) and their receptors correlates with the clinical outcome for lung cancer patients. However, clinical trials of agents that target either of these pathways alone have been disappointing. We hypothesize that targeting both the tumor and its vasculature by simultaneously blocking the VEGFR and EGFR pathways will improve the treatment of locoregional lung cancer. Human lung cancer specimens were first examined for the activation of VEGF receptor 2 (VEGFR2) and EGF receptor (EGFR) for tumor and tumor-associated endothelial cells, and both were found to be activated. The effects of ZD6474 (ZACTIMA), a small-molecule inhibitor of VEGFR2 and EGFR tyrosine kinases, were then studied in vitro using human lung cancer and microvascular endothelial cells. In vitro, ZD6474 inhibited EGFR, VEGFR2, mitogen-activated protein kinase and Akt phosphorylation, EGF- and VEGF-induced proliferation, and endothelial cell tube formation and also induced apoptosis. ZD6474 was further studied in vivo using an orthotopic mouse model of non-small cell lung cancer using NCI-H441 human lung adenocarcinoma cells. The inhibition of both VEGFR2 and EGFR signaling pathways by ZD6474 resulted in profound antiangiogenic, antivascular, and antitumor effects. These results provide a basis for the development of clinical strategies for the combination of selective protein tyrosine kinase inhibitors that block both EGFR and VEGFR signaling as part of the management of locally advanced lung cancer.     

表皮生长因子受体靶向纳米载体荷载C-erbB2反义寡脱氧核苷酸对人乳腺癌SK-BR3细胞的摄取与滞留

背景:结合肿瘤的分子靶向技术和纳米技术,制备出一种新的具有靶向作用的纳米载体,以达到对肿瘤更好靶向作用的目的.目的:制备表皮生长因了偶联牛血清白蛋白纳米载体,联合核素标记的c-erbB2反义寡脱氧核苷酸,体外观察人乳腺癌SK-BR3细胞对其摄取情况.设计、时间及地点:对比观察实验,于2006-09/2008-03在重庆医科大学生物化学和分子生物学教研窀,重庆医科大学分子医学与肿瘤研究中心,重庆医科大学放射医学教研室完成.材料:生血清白蛋白(生物技术级)由美围Amresco公司提供,表皮生长因子由英国PeproTech EC LTD提供,125Ⅰ由成都中核高通同位素股份有限公司提供.寡脱氧核苷酸由上海生工生物工程技术服务有限公司提供.方法:采用超声乳化-化学交联及羧和反应制备表皮生长因子偶联白蛋白靶向纳米载体,通过核素标记示踪技术检测表皮生长因子偶联白蛋白靶向纳米载体的相关性质.主要观察指标:测量表皮生长因子靶向纳米载体荷载c-erbB2反义寡脱氧核苷酸的载药量、包封率及释药率,人乳腺癌SK-BR3细胞对表皮生长因子靶向纳米载体的摄取率及滞留率.结果:表皮生长因子靶向纳米载体包载125Ⅰ标记的反义寡脱氧核苷酸组的摄取率及滞留率高于正义寡脱氧核苷酸组和无义寡脱氧核昔酸组.同时c-erbB2寡脱氧核苷酸使用纳米载体荷载组的摄取率及滞留率均高于未使用纳米载体组,差异有显著性意义(P<0.05).结论:125Ⅰ标记的表皮生长因子靶向纳米载体能够提高乳腺癌SK-BR3细胞对o-erbB2反义寡脱氧核苷酸的摄取和滞留,能达到更好的靶向作用.     

表皮生长因子受体靶向治疗分子影像研究进展

表皮生长因子受体（EGFR）靶向治疗癌症需要准确评估肿瘤EGFR表达。分子影像技术以分子探针与细胞特定靶分子结合，通过PET/CT检测技术获取图像，可无创、准确、重复地实时评价靶分子表达。本文围绕PET各类EGFR分子探针研究进展进行综述。     

基于T2WI影像组学标签预测乳腺癌人表皮生长因子受体2表达状态

目的 探讨基于MR T2WI的影像组学标签在术前预测乳腺癌人表皮生长因子受体2（HER2）表达状态的价值。方法 回顾性收集209例乳腺癌患者的T2WI,将患者随机分为训练组（n=145）和验证组（n=64）。手动勾画病灶ROI,并于Matlab 2013a平台中提取组学特征。通过组间相关系数及最小绝对收缩和选择算子逻辑回归模型筛选组学特征并构建组学标签。比较HER2表达阳性与阴性亚组患者的影像组学得分差异,采用ROC曲线评价训练组中影像组学标签预测HER2的效能,并以获得的预测阈值用于验证组中进行验证。结果 最终获得由13个组学特征构成的影像组学标签。在训练组及验证组中,HER2阳性亚组与阴性亚组患者间组学得分差异均有统计学意义（P均<0.05）。基于T2WI的影像组学标签在训练组及验证组中的AUC分别为0.798、0.707。结论 基于T2WI构建的影像组学标签对术前预测乳腺癌HER2表达状态具有一定价值。     

Expression of transforming growth factor alpha and epidermal growth factor receptor in human bladder cancer.

We analysed the expression of epidermal growth factor receptor (EGFr) and transforming growth factor alpha (TGF-alpha) in human bladder tumours. Tumour biopsies were obtained from 54 patients with primary bladder cancer (18 stage T1 and 36 stage T2-4). The protein and mRNA expression of EGFr and TGF-alpha were quantified by ELISA and competitive RT-PCR, respectively. The EGFr protein level was significantly increased in T2-4 tumours (0.44 x 10(-11); 0.0-27.5 x 10(-11) mol/g) compared with T1 tumours (0.0; 0.0-2.0 x 10(-11) mol/g) (median; range; 2p<0.01). The EGFr protein and mRNA level correlated (Spearman r=0.45, 2p<0.005, n=40). Co-expression of TGF-alpha protein and EGFr protein was significantly associated with muscle invasive tumours (T2-4) (chi-squared=7.9, df=3, p<0.05) and the TGF-alpha protein level correlated significantly with EGFr protein expression (Spearman r=0.56, 2p<0.0001, n=54). While tumour stage correlated with survival, no correlation was observed between survival and the expression of EGFr and/or TGF-alpha. In conclusion, human bladder tumours express both EGFr and TGF-alpha. The expression of EGFr and TGF-alpha are closely correlated, and the expression of EGFr and co-expression of EGFr and TGF-alpha correlate with tumour stage.     

A shift in the treatment of hormone receptor and human epidermal growth factor receptor 2-positive metastatic breast cancer

Historically, postmenopausal women with estrogen receptor (ER)-positive metastatic breast cancer (MBC) with a long disease-free interval and small volume disease have received an aromatase inhibitor. However, the advent of human epidermal growth factor receptor 2 (HER2) testing and its recognition as a poor prognostic indicator has led to the first line use of anti-HER2 directed therapy in combination with chemotherapy. The optimal treatment for those who are both hormone receptor and HER2 receptor positive is less clear. Tumors rich in ER are considered to be less responsive to chemotherapy, and hormone therapy has the benefit of being less toxic than chemotherapy. However, preclinical evidence suggests that HER2 overexpression may confer resistance to endocrine therapy, even in the presence of hormone receptors, due to crosstalk between the two pathways. This review summarizes the evidence from three clinical trials for combining endocrine therapy with anti-HER2 therapy in MBC. The trials raise the possibility of a new treatment approach to co-positive tumors in patients with good performance status and low tumor burden, and a means to potentially delay the need for chemotherapy.     

非小细胞肺癌患者血清人类表皮生长因子受体-2和环氧化酶-2水平及临床意义

目的探讨非小细胞肺癌患者血清人类表皮生长因子受体2（humanepidermalgrowthfactorreceptor2,HER2）和环氧化酶2（cycl00xygenase-2,COX2）水平相关性及与预后的关系。方法非小细胞肺癌患者221例（观察组）与肺部良性疾病患者41例（对照组）,2组采用双抗体夹心ELISA法检测血清HER2和COX2水平,分析二者相关性及与生存时间的关系。结果观察组HER-2和COX-2水平明显高于对照组（P〈0．05）;观察组HER2阳性表达者中位生存时间（48．7周）较HER-2阴性表达者（62．3周）短;COX2阳性表达者中位生存时间（67．0周）较COX-2阴性表达者（83．1周）短（P〈0．05）;观察组HER2与CoX2水平呈正相关（r=0．717,P=0．007）;观察组HER-2和COX-2水平不是影响预后的独立危险因素,但二者联合检测可预测患者预后（OR=1．5,95％CI：1．08～2．09）。结论非小细胞肺癌与HER-2和COX2阳性表达明显相关,二者联合检测有助于非小细胞肺癌的预后评估。     

Effects of the epidermal growth factor receptor inhibitor OSI-774, Tarceva,on downstream signaling pathways and apoptosis in human pancreatic adenocarcinoma

Pancreatic cancer is the fifth leading cause of cancer death in North America. Gemcitabine improves the quality of life of patients but fails to significantly reduce mortality. Our laboratory has demonstrated previously that the phosphatidylinositol 3'-kinase inhibitor wortmannin promotes gemcitabine antitumor activity (S. S. W. Ng et al., Clin. Cancer Res., 7: 3269-3275, 2001). The present study examined the effects of the epidermal growth factor receptor (EGFR) inhibitor OSI-774 ("Tarceva") alone and in combination with wortmannin and/or gemcitabine on downstream signaling molecules, as well as apoptosis in primary pancreatic cancer xenografts implanted orthotopically in severely combined immunodeficient mice. Tumors established from two pancreatic cancer patients [Ontario Cancer Institute Pancreas number (OCIP#) 2 and OCIP#7] were treated with various combinations of the above three drugs and harvested for analyses of the following: the levels of phosphorylated and nonphosphorylated forms of EGFR, protein kinase B (PKB/Akt) and extracellular-regulated kinase (ERK1/2), and the extent of apoptosis using immunofluorescence image analysis and TUNEL assay, respectively. OSI-774 alone significantly inhibited phosphorylation of EGFR in both of the primary xenografts. Phosphorylation of pERK decreased in OCIP#2, but not in OCIP#7. No significant effects on pPKB because of OSI-774 were observed in either tumor type. The extent of apoptosis was significantly increased by 2-fold in OCIP#2 tumors treated with gemcitabine and wortmannin in combination; an additional 2-fold increase in apoptosis was evident in the presence of OSI-774. Although wortmannin failed to enhance gemcitabine-induced apoptosis in OCIP#7 tumors, the extent of apoptosis was significantly increased with the inclusion of OSI-774 in the combination. Taken together, these findings support the use of OSI-774 plus a phosphatidylinositol 3'-kinase inhibitor in combination with gemcitabine in the treatment of pancreatic cancer.     

表皮生长因子受体和血管内皮生长因子在乳腺癌中的表达及临床意义

目的:观察表皮生长因子受体(EGFR)和血管内皮生长因子(VEGF)在乳腺癌组织中的表达特点及临床意义。方法:应用免疫组织化学方法观察EGFR和VEGF在59例乳腺癌组织中的表达情况。结果:59例乳腺癌组织中,EGFR阳性率为93.2%(55/59),VEGF染色全部呈阳性(59/59),EGFR表达在乳腺癌组织分级和淋巴结是否转移之间比较差异有显著性(均P<0.05)。PR阳性组中VEGF表达水平较高(P<0.05),EGFR和VEGF两者的表达呈正相关(P<0.05)。结论:EGFR和VEGF在乳腺癌的生物学行为中起着一定的作用,研究两者联合表达有一定的临床意义。