药学学报

Items	Contents for definition of acquired resistance to EGFR TKIs in lung cancer
1	Previously administered a single-agent EGFR TKI (eg, gefitinib or erlotinib) for treatment
	One of the following: A. A well-known EGFR mutation that is related to drug sensitivity (eg, G791X, exon 19 deletion, L858R, L861Q) B. Real clinical benefit from therapy of an EGFR TKI as defined by one of the following: ⅰ Efficacy evaluation of partial or complete response (RECIST or WHO) ⅱ Statistically meaningful and lasting clinical benefits (stable state of lasting ≥ 6 months after initial treatment, as defined by RECIST or WHO)
2	Continuous therapy of gefitinib or erlotinib for the last 30 days after systemic progression of disease
3	No intervening any treatment during termination of gefitinib or erlotinib and start of new drug treatment

Items	Contents for definition of acquired resistance to EGFR TKIs in lung cancer
1	Previously administered a single-agent EGFR TKI (eg, gefitinib or erlotinib) for treatment
	One of the following: A. A well-known EGFR mutation that is related to drug sensitivity (eg, G791X, exon 19 deletion, L858R, L861Q) B. Real clinical benefit from therapy of an EGFR TKI as defined by one of the following: ⅰ Efficacy evaluation of partial or complete response (RECIST or WHO) ⅱ Statistically meaningful and lasting clinical benefits (stable state of lasting ≥ 6 months after initial treatment, as defined by RECIST or WHO)
2	Continuous therapy of gefitinib or erlotinib for the last 30 days after systemic progression of disease
3	No intervening any treatment during termination of gefitinib or erlotinib and start of new drug treatment

Group	First generation	Second generation	Third generation
Mutation sites	L858R, del E746_A750	L718Q, L844V and T790M	C797S, A768_V769dupASV
Drugs	Gefitinib, erlotinib	Afatinib, neratinib, CL-387, 785	AZD9291, CO-1686, WZ400, HM61713
Target sites	del E746_A750 or L858R/L718Q, L844V or C797S	del E746_A750 or L858R/L718Q, L844V or C797S; Q791R; wild type (afatinib)	T790M; L718Q, L844V and L858R /L844V (AZD9291)

Group	First generation	Second generation	Third generation
Mutation sites	L858R, del E746_A750	L718Q, L844V and T790M	C797S, A768_V769dupASV
Drugs	Gefitinib, erlotinib	Afatinib, neratinib, CL-387, 785	AZD9291, CO-1686, WZ400, HM61713
Target sites	del E746_A750 or L858R/L718Q, L844V or C797S	del E746_A750 or L858R/L718Q, L844V or C797S; Q791R; wild type (afatinib)	T790M; L718Q, L844V and L858R /L844V (AZD9291)

非小细胞肺癌治疗药物EGFR-TKIs获得性耐药机制的研究进展

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王晨 , 陈淑珍 ^*

药学学报 | 综述 2019,54(8): 1364-1371

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药学学报 | 综述 2019, 54(8): 1364-1371

非小细胞肺癌治疗药物EGFR-TKIs获得性耐药机制的研究进展

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王晨, 陈淑珍^*

作者信息

中国医学科学院、北京协和医学院医药生物技术研究所, 北京 100050

通讯作者:

*陈淑珍, Tel:86-10-63166305, Fax:86-10-63138137, E-mail:bjcsz@imb.pumc.edu.cn

Advances in the mechanisms of acquired resistance to EGFR-tyrosine kinase inhibitors in non-small cell lung cancer

Chen WANG, Shu-zhen CHEN^*

Affiliations

Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, China

出版时间: 2019-08-12 doi: 10.16438/j.0513-4870.2019-0089

文章导航

摘要

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肺癌在所有肿瘤中死亡率占第一位，其中非小细胞肺癌（NSCLC）占肺癌发病率的80%左右。近年来，分子靶向药物发展迅速，其中表皮生长因子受体酪氨酸激酶抑制剂（EGFR-TKIs）以表皮生长因子受体（EGFR）作为靶点，在治疗NSCLC患者中取得很好的疗效，目前有三代EGFR-TKIs在临床上使用，效果优于传统化疗，但是也都出现获得性耐药的问题。本文将获得性耐药机制的研究进展分为依赖EGFR通路和非依赖EGFR通路两部分作一综述。依赖EGFR通路主要介绍EGFR的基因突变，非依赖EGFR通路包括HER2的扩增、BIM缺失、HGF/c-MET通路的激活、IGF1R的活化、RAS突变、PTEN缺失、上皮-间充质转化、PUMA表达下调、IL-6和VEGF高表达，这些耐药机制相互联系，相互影响，使肿瘤细胞对EGFR-TKIs产生耐药。

关键词

非小细胞肺癌 / 表皮生长因子受体酪氨酸激酶抑制剂 / 获得性耐药 / 表皮生长因子受体依赖性 / 表皮生长因子受体非依赖性

Abstract

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Lung cancer ranks the first in the mortality rate among all cancers, and non-small cell lung carcinoma (NSCLC) accounts for about 80% of the incidence of lung cancer. In recent years, the drugs targeting specific molecules have been developed rapidly. The epidermal growth factor receptor-tyrosine kinase inhibitors (EGFRTKIs) have achieved good results in the treatment of patients with NSCLC. Currently, there are three generations of EGFR-TKIs, and the treatment outcome of these drugs surpasses traditional chemotherapies. However, the problems of acquired resistance remains in the course of treatment. In this review, research progress of the mechanisms of acquired resistance is divided into two parts:EGFR-dependent pathway and EGFR-independent pathway. The EGFR-dependent pathway mainly includes EGFR gene mutations, whereas the EGFR-independent pathways include HER2 amplification, BIM deletion, activation of HGF/c-Met pathway, activation of IGF1R, RAS mutation, PTEN deletion, epithelial-mesenchymal transition, PUMA loss, and high levels of expression of VEGF or IL-6. These interconnected mechanisms are linked with acquired resistance to EGFR-TKIs in NSCLC.

Key words

non-small cell lung cancer / epidermal growth factor receptor-tyrosine kinase inhibitor / acquired resistance / EGFR-dependence / EGFR-independence

引用本文

王晨, 陈淑珍. 非小细胞肺癌治疗药物EGFR-TKIs获得性耐药机制的研究进展. 药学学报, 2019 , 54 (8) : 1364 -1371 . DOI: 10.16438/j.0513-4870.2019-0089

Chen WANG, Shu-zhen CHEN. Advances in the mechanisms of acquired resistance to EGFR-tyrosine kinase inhibitors in non-small cell lung cancer[J]. Acta Pharmaceutica Sinica, 2019 , 54 (8) : 1364 -1371 . DOI: 10.16438/j.0513-4870.2019-0089

正文

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肺癌在全世界范围内的发病率和死亡率逐年上升, 其中约80%是非小细胞肺癌(non-small cell lung cancer, NSCLC), 表皮生长因子受体(epidermal growth factor receptor, EGFR)的激活突变是NSCLC的关键驱动因素^[1]。靶向EGFR的表皮生长因子受体酪氨酸激酶抑制剂(epidermal growth factor receptor tyrosine kinase inhibitor, EGFR-TKIs)能够有效治疗EGFR突变阳性的NSCLC。目前, 已有三代EGFR-TKIs在临床上使用, 第一代EGFR-TKIs有厄洛替尼、吉非替尼等, 作用于EGFR胞内蛋白酪氨酸激酶, 与ATP可逆性竞争结合于酪氨酸激酶功能域^[2], 抑制其磷酸化, 目前主要用于激活突变的晚期NSCLC患者。第二代EGFR-TKIs, 如阿法替尼。与第一代EGFR-TKIs不同的是, 阿法替尼不可逆性竞争地结合于酪氨酸激酶功能域^[3], 抑制其磷酸化, 对野生型EGFR也有抑制作用。第三代EGFR-TKIs如AZD9291, 也可以不可逆地抑制酪氨酸激酶磷酸化, 并且对T790M耐药突变的患者有较好的治疗效果^[4]。但是, 在使用第一代EGFR-TKIs治疗9~12月后, 患者出现了获得性耐药^[5]。60%患者因为EGFR在T790M上的突变而耐药^[6]。第二代不可逆的EGFR-TKIs阿法替尼对T790M突变型的患者也没有好的疗效, 针对T790M突变的第三代EGFR-TKIs对其有着较好的疗效, 可是会再次出现耐药。2010年1月由哈佛医学院Jackman等^[7]共同讨论制定了NSCLC患者EGFR-TKIs获得性耐药的定义, 见表 1。目前的研究发现, 除了EGFR突变导致的获得性耐药, 还有一些别的耐药机制, 本文就EGFR-TKIs获得性耐药机制分为依赖EGFR通路和非依赖EGFR通路进行论述。

1 依赖EGFR通路

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随着EGFR-TKIs的发展, EGFR共发生3次相应的突变。第1次为EGFR激活突变, 最常见为外显子21的Leu858Arg (L858R)突变和外显子19的delE746-A750缺失^{[8, 9]}, 对第一代或第二代EGFR酪氨酸激酶抑制剂如厄洛替尼、吉非替尼或阿法替尼的治疗有良好的反应^[10]。但是, 经过一段时间的起初治疗后, 60%患者出现了获得性耐药, 主要因为EGFR第2个位点的突变Thr790Met (T790M), 即苏氨酸变成甲硫氨酸; 而非依赖EGFR途径, 如MET和HER2的扩增、EGFR的扩增、PIK3CA的突变等^[11]情况出现得较少。密码子790 (T790M)中的苏氨酸到甲硫氨酸的变化发生在酪氨酸激酶域, 增加ATP亲和力, 从而阻止第一代可逆EGFR-TKIs与EGFR酪氨酸激酶域的结合^[8]。

第三代EGFR-TKIs, 例如AZD9291、HM61713、CO-1686在起始剂量下就可以对T790M阳性的患者产生很好的疗效, 但一些患者在用药几个月后又出现了疾病进展, Ercan等^[12]发现, 耐药株出现了EGFR L718Q、L844V和C797S 3种突变。在C797S、del 19或L858R和T790M存在的情况下, 患者会对现有EGFR抑制剂产生抵抗。Thress等^[13]发现C797S突变与T790M突变都在20号外显子上, 可使其797位点上的半胱氨酸被丝氨酸所取代, 导致第三代EGFR-TKIs无法与EGFR蛋白发生共价结合, 从而发生耐药。他们进一步对C797S突变序列做了研究。在对编号为1的发生C797S突变的耐药患者的血清进行靶向测序, 鉴定出获得性T→A突变, 其编码EGFR C797S突变, 并且C797S和T790M突变在同一等位基因上呈顺式关系。而对2号患者的血清测序中发现, 其C797S突变为G→C突变, 且与T790M在不同的等位基因上呈反式关系。接着, 4号、5号患者的肿瘤活检和血清测序同时发现, 两位患者在这2个位点上均发生C797S突变, 但同时发生T→A和G→C突变, 所以, 同样发生C797S突变, 突变位点也有所不同。同时, 他们也发现, 在第三代EGFR-TKIs的使用下, 只有一部分患者发生了C797S突变, 他们对15位使用AZD9291的患者的样本做了数字PCR, 发现治疗前, T790M均为阳性, 但在耐药时出现3个分子亚型: 6例获得C797S突变, 5例维持T790M突变, 但未获得C797S突变, 4例检测到EGFR激活突变而丢失T790M突变。这说明C797S突变只是患者对第三代EGFR-TKIs耐药的主要原因之一。

患者对EGFR-TKIs的耐药原因不一, 如对三代EGFR-TKIs WZ4002、CO-1686和AZD9291来说, EGFR同时发生L718Q、L844V和C797S突变时, 对WZ4002和CO-1686产生耐药。但仅C797S突变就会引起患者对AZD9291耐药。另外, 一个EGFR-TKIs并不是只对某一种基因型的肿瘤细胞起效, 第一代EGFR-TKIs吉非替尼对EGFR第一次激活突变(L858R和del E746_A750)的细胞敏感, 同时对于有着突变del E746_ A750/C797S和二代突变L844V的细胞也保持敏感性。阿法替尼对于EGFR野生型和激活突变的细胞均有很好的敏感性, 同时对于Q791R (克隆2372, A > G)突变、二代突变L718Q和L844V以及del E746_A750/C797S突变细胞有敏感性^[9]。这为治疗三代EGFR-TKIs耐药提供思路, 出现C797S突变, 在T790M阴性的情况下, 重新启用吉非替尼和阿法替尼可以带来很好的获益。EGFR三代突变的位点, 相应代表药物及其作用靶点的总结见表 2。

2 非依赖EGFR通路

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2.1 HER2扩增

人类表皮生长因子受体2 (human epidermal growth factor receptor-2, HER2)基因, 即c-ErbB-2基因, 和EGFR均属于ErbB受体家族的一种, 该家族包括EGFR (ErbB-1)、HER2/c-neu (ErbB-2)、HER3 (ErbB-3)和HER4 (ErbB-4)。HER2为原癌基因^[14], 是一种酪氨酸激酶受体, 研究表明HER2在调控肿瘤细胞对EGFR-TKIs的耐药中起重要作用, HER2扩增和EGFR T790M突变一般不同时出现。Planchard等^[15]在研究2个服用AZD9291后耐药的病例中发现, HER2为第三代EGFR-TKIs获得性抵抗的潜在机制, 并且扩增出现在T790M突变完全消失的时候。他们推测是由于在AZD9291治疗开始时, 药物抑制了T790M阳性细胞的增长, 即在肿瘤细胞中存在对T790M阴性克隆的选择, T790M阴性细胞可以增长, 此时, 其耐药性源于非依赖EGFR途径, 即HER2扩增。所以, 使用AZD9291发生耐药时, HER2扩增可能成为了耐药的主要原因, 若将EGFR-TKIs和HER2抑制剂联用可能带来更好的临床治疗效果。

2.2 Bcl-2蛋白家族促凋亡介质(Bcl-2 interacting mediator of cell death, BIM)

BIM缺失多态性是可以预测EGFR-TKIs治疗效果的标志物, BIM基因上大约2 900 bp的缺失多态性会下调BIM蛋白的促凋亡功能, 导致了NSCLC细胞的耐药^[16]。一项对871位NSCLC患者的meta分析结果表明, 对于EGFR基因突变的NSCLC患者, 存在BIM基因缺失多样性的患者EGFR-TKIs治疗效果不佳, 疾病控制率和无进展生存期等都减少, BIM基因缺失多样性可能导致EGFR-TKIs耐药^[17]。伏立诺他和二甲双胍联用可以上调BIM的表达, 进而改善耐药细胞对吉非替尼的敏感性^[18], 因此, 以BIM为靶点, 上调BIM的表达可以成为改善细胞耐药的新方法。

2.3 HGF/c-MET通路

MET为一种原癌基因, 也是跨膜络氨酸激酶受体, 激活后通过下游ERK、AKT、STATs通路影响细胞增殖和凋亡水平。近期研究表明, 导致肿瘤细胞对EGFR-TKIs耐药的原因中, 有很多都与MET扩增及其下游PI3K/AKT通路的过度激活有关, 如一些膜蛋白、核苷酸和激酶的表达异常, 导致下游PI3K/AKT通路激活, 进而又通过激活下游信号转导促进肿瘤发展^[19]。MET扩增与EGFR发生T790M突变在导致药物耐药中是相互独立或者互补的关系。MET扩增是通过加强ErbB3的磷酸化和下调PI3K/AKT通路来诱导一代EGFR-TKIs的耐药^[20]。Shi等^[21]在联用三代药物AZD9291和PF02341066 (MET抑制剂)对HCC827/ER和HCC827/AR进行体内外实验时得出结论, 药物的联用下调了ErbB3/ERK通路, 进而抑制了BIM的磷酸化, 使BIM稳定从而诱导细胞凋亡, 产生很好的抑制肿瘤细胞增殖的效果。可见MET扩增在导致药物耐药中起到重要作用。肝细胞生长因子(hepatocyte growth factor, HGF)通过影响MET基因在肿瘤发生发展和肿瘤耐药中扮演着重要作用。Yang等^[22]研究发现, 在EGFR野生型的肺癌患者中, HGF可以通过MET或者下游PI3K通路, 降低患者对吉非替尼的敏感性, 但是在使用HGF抑制性单克隆抗体、MET抑制剂或者MET敲除的处理后, 敏感性得以恢复。Jang等^[23]发现, 腺苷酸转移酶2 (adenylate transferase 2, ANT2)为真核细胞线粒体内膜上转运蛋白家族成员之一, 它的过表达会导致肿瘤细胞对EGFR-TKIs耐药。在对非小细胞肺癌细胞H1975和HCC827/GR细胞的ANT2基因敲除之后, Hsp90在细胞中的表达受到抑制, PI3K/AKT通路活性下调, 从而克服了细胞对EGFR-TKIs的耐药, 最终使细胞生长受到抑制。Deng等^[24]研究了PI3K/AKT通路在吉非替尼耐药中的作用, 结果表明, 相对于PC9细胞, PC9/G (吉非替尼耐药)细胞中AKT的磷酸化水平显著升高, ERK的磷酸化水平却相近; 同时, DNA水平分析表明, 这2种细胞之间只有整合素β1的表达水平显著不同, 因而整合素β1引起的吉非替尼耐药是由PI3K/AKT通路引起的。随后, 敲低整合素β1减少其表达或者使用PI3K的抑制剂LY294002, AKT的磷酸化都受到抑制, 耐药细胞对吉非替尼的敏感性、凋亡水平、G₀/G₁的比例都有所上升。HGF/c-MET通路及其下游ERK、PI3K/AKT、STATs通路在NSCLC细胞对EGFR-TKIs的耐药中产生重要影响, 使用抑制剂干预通路上某一蛋白的表达, 或者对某一蛋白的基因进行敲低, 可以改善其耐药。

2.4 胰岛素样生长因子1受体(insulin-like growth factor 1 receptor, IGF1R)

IGF1R是人类细胞表面蛋白质, 属于络氨酸激酶受体家族, 其配体是胰岛素样生长因子-1 (insulin-like growth factor 1, IGF-1)。Yi等^[25]发现, HGF和IGF-1的分泌增加膜联蛋白A2 (annexin A2, ANXA2)的表达和磷酸化, 通过双抑制HGF/c-MET和IGF-1/IGF-1R通路可显著抑制ANXA2, 从而显著降低肿瘤上皮-间充质转化(epithelial-mesenchymal transition, EMT)和吉非替尼的耐药。Chen等^[26]在对核苷酸miR-223在EGFR-TKIs耐药中起到的作用的研究发现, miR-223的表达上调可以导致IGF1R的mRNA及蛋白水平表达的下调, 进而抑制下游PI3K/AKT/mTOR通路, 最终可以改善肿瘤细胞对EGFR-TKIs的敏感性。Suda等^[27]使用EGFR野生型的H358细胞构建厄洛替尼的耐药株, 他们发现H358/ER细胞没有产生EGFR的二代突变, 也没有发现MET基因的扩增, 但是他们发现细胞内IGF1R磷酸化水平升高了, 因此他们推断, IGF1R的活化是导致EGFR野生型肺癌细胞对厄洛替尼耐药的机制之一。Guerard等^[28]在对IGF1R导致细胞对吉非替尼耐药的机制的深入研究中发现了一种生长因子双调蛋白, 此调节蛋白可以使IGF1R从胞外向胞核转移, 而胞核内IGF1R的累积, 可以引起细胞周期的阻滞, 因此阻碍细胞发生凋亡, 产生对吉非替尼的耐药。IGF1R的活化可以通过激活下游PI3K/AKT/mTOR通路或上调膜联蛋白A2的表达等来下调细胞的凋亡水平并促使细胞产生耐药。

2.5 RAS突变

在非小细胞肺癌的患者中, 西方和亚洲人群分别有26%和11%的患者出现k-RAS基因上的突变并都有不良预后^[29]。Ohba等^[30]在一项回顾性分析中指出, 在354例已手术切除的肺腺癌患者中, EGFR和RAS基因突变分别发生在41.1%和6.4%的患者中, 虽然RAS基因突变的比例不高, 但是RAS突变阳性的患者比阴性患者的无进展生存率和总体生存率都要低很多, 其中5年内无进展生存率是50.8%比76.9%, 总体生存率是70.0%比86.6%, 表明RAS基因突变是一个预后不良的独立预测因素。Zhou等^[31]在对45名服用吉非替尼的非小细胞肺癌患者的回顾性分析中发现, 45名患者中有8名出现了k-RAS基因的突变, 突变发生在k-RAS基因1号和2号外显子的12位、13位密码子上。发生突变的患者与未发生的患者的客观缓解率有显著性差异。所以k-RAS基因突变与否对吉非替尼的临床获益产生较大影响。Yamaoka等^[32]构建了阿法替尼的耐药株, 他们发现有些耐药细胞的野生型k-RAS基因发生过表达情况, 这导致了EGFR和下游信号分子ERK1/2或AKT之间信号转导发生障碍。在撤药2个月之后, k-RAS的表达恢复母本细胞水平, 肿瘤细胞也恢复了对阿法替尼的敏感性。k-RAS基因的突变情况可以作为细胞对EGFR-TKIs的敏感性的独立预测因素。

2.6 PTEN缺失

磷酸酶和张力蛋白类似物(phosphatase and tensin homolog, PTEN)是一种抑癌基因, 抑制其表达会促进恶性肿瘤的增长^[33]。Sos等^[34]在EGFR依赖性的肺癌细胞中发现, PTEN缺失可以部分使EGFR的突变与下游信号传导解偶联并激活EGFR, 导致细胞对厄洛替尼耐药。PTEN通过去磷酸化磷脂酰肌醇-3, 4, 5-三磷酸(phosphatidylinositol-3, 4, 5-triphosphate, PIP3)来抵消AKT活化, 因此PTEN缺失也可以使AKT活化, 导致细胞耐药。过氧化物酶体增殖物激活受体γ^[35] (peroxisome proliferator-activated receptor gamma, PPARγ)是一种核受体, 其被激活后可上调PTEN表达进而抑制PI3K下游的细胞信号传导, 介导细胞凋亡。所以, PTEN的缺失会导致NSCLC细胞对EGFR-TKIs产生耐药, 上调其表达为改善EGFR-TKIs的耐药提供了新的思路。

2.7 EMT

EMT是指具有极性的上皮细胞丧失极性, 转化成具有迁移能力的间质细胞的过程^[36]。肿瘤细胞出现EMT表型后进入了癌症干细胞状态, 此时, 常规治疗通常不能根治^[37]。Li等^[38]发现, 与吉非替尼敏感的亲代细胞相比, 具有EMT表型的吉非替尼耐药细胞显示出更强的迁移和侵袭能力。逆转吉非替尼耐药细胞的EMT表型, 可以部分恢复对吉非替尼的敏感性。TWIST1是EMT转录因子, 基因沉默或TWIST1抑制剂可以抑制EGFR突变的NSCLC细胞生长^[39]。TWIST1的过表达导致NSCLC患者对厄洛替尼和AZD9291的耐药。BIM是促凋亡BH3基因, TWIST1通过直接结合其内含子区和启动子而抑制BIM的转录。所以, EMT的出现可以导致患者对EGFR-TKIs产生耐药, 此时EMT转录因子TWIST1的过表达抑制耐药细胞凋亡基因的表达, 对药物产生抵抗。

2.8 p53上调的细胞凋亡调节剂(p53 upregulated modulator of apoptosis, PUMA)

PUMA是Bcl-2家族中唯BH3域蛋白, 在p53引起的凋亡中是必需的^[40]。PUMA与EGFR-TKIs对肿瘤的敏感性有关, PUMA的活性增强可以增加吉非替尼引起的肿瘤细胞凋亡; 相反, 使用shRNA抑制PUMA的表达, 可以削弱吉非替尼的作用。Han等^[41]通过比较H1650吉非替尼耐药和非耐药细胞的转录组和基因组的改变, 发现耐药细胞中的促凋亡基因PUMA的表达下调了。PUMA的丢失或突变会导致NSCLC细胞对EGFR-TKIs的耐药, 增强其表达可以增加肿瘤细胞凋亡水平, 进而提高对肿瘤细胞的杀伤作用。

2.9 旁路途径

一些细胞因子活化或激酶激活的旁路途径, 如白细胞介素-6的活化(interleukin-6, IL-6)和血管内皮生长因子的高表达(vascular endothelial growth factor, VEGF)等, 可以导致肿瘤细胞对药物的耐药。肿瘤细胞中IL-6的高表达预示着EGFR突变的NSCLC患者对吉非替尼的获益减小^[42]。一项对52名接受过吉非替尼治疗的EGFR突变的NSCLC患者的回顾性分析^[43]表明, IL-6高表达的患者无进展生存期更短。对于铂类化疗方式治疗的患者, IL-6的表达与否对患者无进展生存期没有影响, 说明IL-6主要影响用EGFR-TKIs治疗患者的效果。联用EGFR-TKIs和抗IL-6的抗体, 可以增加EGFR突变的NSCLC患者对EGFR-TKIs的敏感性。由于VEGF与EGFR有很多相互联系, 因此高度选择性的血管内皮生长因子2 (vascular endothelial growth factor receptor 2, VEGFR2)抑制剂阿帕替尼可以显著地强化吉非替尼对T790M阳性突变细胞的抗肿瘤效果^[44]。人参皂苷Rg3可以下调VEGF和VEGFR的表达, 同样可以改善患者对EGFR-TKIs的耐药, 延长患者无进展生存期^[45]。IL-6和VEGF, 因其与EGFR可以直接相互影响, 所以它们的高表达会导致EGFR突变的NSCLC患者对EGFR-TKIs的耐药。

2.10 程序化死亡配体(programmed death ligand, PD-L1)

PD-L1与肿瘤免疫逃逸和预后不良有关, 它是否影响EGFR突变型NSCLC患者对EGFR-TKIs的耐药还存在争议, 目前还没有一致的结论。Tung等^[46]预测, 不管NSCLC患者EGFR基因突变与否, PD-L1的高表达可以使细胞对EGFR-TKIs产生耐药。通过对PD-L1进行基因改造, 发现缺氧诱导因子-1α (hypoxia inducible factor-1α, HIF-1α)和YES关联蛋白1 (YES associated protein 1, YAP1)表达的变化与PD-L1的表达变化存在相关性。PD-L1沉默可以使HIF-1α和YAP1的表达下调。其中, PD-L1调节的YAP1表达量的变化可以改变吉非替尼杀伤肿瘤细胞的IC₅₀值, 而且在耐药患者中, 通常PD-L1和YAP1的mRNA表达同时升高, 所以推测被PD-L1调控的YAP1表达上调可能导致EGFR-TKIs的耐药。Kunimasa等^[47]在分析临床样本时发现, 若患者肿瘤细胞中PD-L1高表达, 临床疾病进程较快。对于PD-L1高表达, 没有T790M突变的EGFR突变型NSCLC患者, PD-1抑制治疗有更好的效果。Su等^[48]在对101位受试者的研究中发现, PD-L1的高表达会显著降低患者的客观缓解率和无进展生存期, PD-L1的表达导致细胞对EGFR-TKIs的耐药。另外, PD-L1的阳性表达主要出现在对EGFR-TKIs先天耐药的患者中, 而不是获得性耐药的患者, 对于没有发生获得性耐药突变的耐药患者, 他们的研究提供了PD-1阻断以治疗耐药的新思路。Kobayashi等^[49]对211例NSCLC患者样本的回顾性分析中得出结论, PD-L1阳性患者的总体生存率比PD-L1阴性患者低, 但是对于携带EGFR突变的NSCLC患者, PD-L1阳性与否对EGFR-TKIs的效果没有影响。Suda等^[50]研究了靶向PD-1或PD-L1的治疗对于EGFR突变NSCLC患者效果不好的原因, 结果发现, EGFR-TKIs获得性耐药细胞的EGFR磷酸化水平升高, 进而上调PD-L1的表达, 但是非耐药突变的细胞没有出现PD-L1高表达的情况。所以, PD-1或PD-L1的表达情况可能只对于发生耐药突变的NSCLC患者有治疗上的指导意义。有关PD-L1在EGFR-TKIs耐药中的作用需要针对具体情况进行分析。

3 总结与展望

收起

靶向EGFR的EGFR-TKIs在临床上产生了很好的疗效, 但是随着EGFR的不断突变, EGFR-TKIs都出现耐药的问题。同时, 非依赖EGFR通路, 如HER2的扩增、BIM缺失、HGF/c-MET通路的激活、IGF1R的活化、RAS突变、PTEN缺失、EMT、PUMA表达下调、IL-6、VEGF高表达等, 都可能导致非小细胞肺癌细胞对EGFR-TKIs产生耐药(图 1)。某一通路的改变也不只有一个效应分子, 如HGF/c-MET通路的激活会影响其下游ERK、PI3K/AKT、STATs通路, IGF1R的活化除了激活下游PI3K/AKT/mTOR通路, 还可以增加膜联蛋白A2的表达。不同耐药机制之间相互联系, 相互影响。如HGF/c-MET通路的激活、IGF1R的活化和PTEN缺失都会影响下游PI3K/AKT通路, 进而对细胞凋亡水平产生影响。EMT的转录因子TWIST1会抑制BIM的转录, 共同促使细胞产生耐药。非小细胞肺癌对EGFR-TKIs产生获得性耐药的机制复杂, 可能还存在尚未阐明的耐药机制, 同时, 各种耐药机制之间的相互调节也需要进一步研究, 针对不同患者的耐药机制采取独特的治疗方案即个体化治疗将是克服耐药的主要研究方向。

随着细胞和分子生物学技术的发展, NSCLC对EGFR-TKIs耐药的分子机制逐渐阐明, 因而, 研究者能够以此为基础, 建立依据多个通路的筛选模型, 发现活性化合物, 并对其进行结构修饰, 再进行体内外药效学、药代动力学、毒性实验评价, 从而发现更多的候选药物并推向临床, 克服EGFR-TKIs耐药, 提高NSCLC患者的生存率; 同时耐药机制的阐明, 带来一些EGFR-TKIs联合其他治疗的手段, 例如EGFR-TKIs联合MET抑制剂、EMT和PI3K/AKT通路小分子抑制剂、上调BIM和PUMA小分子调节剂、IGF1R单克隆抗体、MET基因敲除、IL-6和VEGF的抑制剂等。但是, 寻求更好的联合用药方式, 延长患者生存期, 提高患者生存质量, 仍需进一步研究。

基金

收起

国家自然科学基金资助项目(81621064)
中国医学科学院医学与健康科技创新工程(2016-I2M-2-002)
重大新药创制科技重大专项(2018ZX09711001-007-002)

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2019年第54卷第8期

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doi: 10.16438/j.0513-4870.2019-0089

接收时间：2019-01-24
首发时间：2026-01-26
出版时间：2019-08-12

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收稿日期：2019-01-24
修回日期：2019-03-15

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国家自然科学基金资助项目(81621064)

中国医学科学院医学与健康科技创新工程(2016-I2M-2-002)

重大新药创制科技重大专项(2018ZX09711001-007-002)

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中国医学科学院、北京协和医学院医药生物技术研究所, 北京 100050

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*陈淑珍, Tel:86-10-63166305, Fax:86-10-63138137, E-mail:bjcsz@imb.pumc.edu.cn

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2种不同金属材料的力学参数

科 Family	属数 Number of genus	种数 Number of species	占总种数比例 Percentage of total species (%)	属 Genus	种数 Number of species	占总种数比例 Percentage of total species (%)
鹅膏菌科Amanitaceae	2	11	5.26	鹅膏菌属 Amanita	10	4.78
小菇科 Mycenaceae	2	12	5.74	丝盖伞属 Inocybe	5	2.39
多孔菌科 Polyporaceae	8	14	6.70	蜡蘑属 Laccaria	5	2.39
红菇科 Russulaceae	3	23	11.00	小皮伞属 Marasmius	6	2.87
				小菇属 Mycena	11	5.26
				光柄菇属 Pluteus	5	2.39
				红菇属 Russula	17	8.13
				栓菌属 Trametes	5	2.39

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Items	Contents for definition of acquired resistance to EGFR TKIs in lung cancer
1	Previously administered a single-agent EGFR TKI (eg, gefitinib or erlotinib) for treatment
	One of the following: A. A well-known EGFR mutation that is related to drug sensitivity (eg, G791X, exon 19 deletion, L858R, L861Q) B. Real clinical benefit from therapy of an EGFR TKI as defined by one of the following: ⅰ Efficacy evaluation of partial or complete response (RECIST or WHO) ⅱ Statistically meaningful and lasting clinical benefits (stable state of lasting ≥ 6 months after initial treatment, as defined by RECIST or WHO)
2	Continuous therapy of gefitinib or erlotinib for the last 30 days after systemic progression of disease
3	No intervening any treatment during termination of gefitinib or erlotinib and start of new drug treatment

Items

Contents for definition of acquired resistance to EGFR TKIs in lung cancer

Previously administered a single-agent EGFR TKI (eg, gefitinib or erlotinib) for treatment

One of the following:
  A. A well-known EGFR mutation that is related to drug sensitivity (eg, G791X, exon 19 deletion, L858R, L861Q)
  B. Real clinical benefit from therapy of an EGFR TKI as defined by one of the following:
    ⅰ Efficacy evaluation of partial or complete response (RECIST or WHO)
    ⅱ Statistically meaningful and lasting clinical benefits (stable state of lasting ≥ 6 months after initial treatment, as defined by RECIST
      or WHO)

Continuous therapy of gefitinib or erlotinib for the last 30 days after systemic progression of disease

No intervening any treatment during termination of gefitinib or erlotinib and start of new drug treatment

Group	First generation	Second generation	Third generation
Mutation sites	L858R, del E746_A750	L718Q, L844V and T790M	C797S, A768_V769dupASV
Drugs	Gefitinib, erlotinib	Afatinib, neratinib, CL-387, 785	AZD9291, CO-1686, WZ400, HM61713
Target sites	del E746_A750 or L858R/L718Q, L844V or C797S	del E746_A750 or L858R/L718Q, L844V or C797S; Q791R; wild type (afatinib)	T790M; L718Q, L844V and L858R /L844V (AZD9291)

Group

First generation

Second generation

Third generation

Mutation sites

L858R, del E746_A750

L718Q, L844V and T790M

C797S, A768_V769dupASV

Drugs

Gefitinib, erlotinib

Afatinib, neratinib, CL-387, 785

AZD9291, CO-1686, WZ400, HM61713

Target sites

del E746_A750 or L858R/L718Q,
L844V or C797S

del E746_A750 or L858R/L718Q, L844V
or C797S; Q791R; wild type (afatinib)

T790M; L718Q, L844V and L858R
/L844V (AZD9291)