The epidermal growth factor receptor (EGFR) is a tyrosine kinase receptor that plays an important role in normal epidermal cell physiology. EGFR is overexpressed in cancer cells and has a number of mutations that implicate tumor malignancy, development, and poor patient prognosis; thus, EGFR is an attractive target for cancer therapy. At present, anti-EGFR monoclonal antibodies (mAbs) have been approved and are used for treating patients with a variety of EGFR-expressing cancers. Epitope mapping is important in identifying the therapeutic mechanism of anti-EGFR mAbs; however, the development of epitope mapping techniques lags behind the development of antimolecular target mAbs, including anti-EGFR mAbs. Hence, in this study, a novel epitope mapping method, RIEDL insertion for epitope mapping (REMAP) method, was developed. The results of this study demonstrated that the critical epitope of anti-EGFR mAb EMab-134 is Gly378, Asp379, Ser380, Phe381, Thr382, His383, Thr384, Pro385, and Pro386 of EGFR. The REMAP method could be useful for determining the critical epitope of functional mAbs against many target molecules.
Get full access to this article
View all access options for this article.
References
1.
OlayioyeMA, NeveRM, LaneHA, and HynesNE: The ErbB signaling network: Receptor heterodimerization in development and cancer. EMBO J, 2000; 19:3159–3167.
2.
PinesG, KöstlerWJ, and YardenY: Oncogenic mutant forms of EGFR: Lessons in signal transduction and targets for cancer therapy. FEBS Lett, 2010; 584:2699–2706.
3.
LewisTS, ShapiroPS, and AhnNG: Signal transduction through MAP kinase cascades. Adv Cancer Res, 1998; 74:49–139.
GschwindA, FischerOM, and UllrichA: The discovery of receptor tyrosine kinases: Targets for cancer therapy. Nat Rev Cancer, 2004; 4:361–370.
6.
FanQW, ChengC, KnightZA, Haas-KoganD, StokoeD, JamesCD, McCormickF, ShokatKM, and WeissWA: EGFR signals to mTOR through PKC and independently of Akt in glioma. Sci Signal, 2009; 2:ra4.
7.
SigismundS, AvanzatoD, and LanzettiL: Emerging functions of the EGFR in cancer. Mol Oncol, 2018; 12:3–20.
8.
ArataniK, KomatsuS, IchikawaD, OhashiT, MiyamaeM, OkajimaW, ImamuraT, KiuchiJ, NishibeppuK, KosugaT, KonishiH, ShiozakiA, FujiwaraH, OkamotoK, TsudaH, and OtsujiE: Overexpression of EGFR as an independent prognostic factor in adenocarcinoma of the esophagogastric junction. Anticancer Res, 2017; 37:3129–3135.
9.
NanX, XieC, YuX, and LiuJ: EGFR TKI as first-line treatment for patients with advanced EGFR mutation-positive non-small-cell lung cancer. Oncotarget, 2017; 8:75712–75726.
10.
IsomotoK, HarataniK, HayashiH, ShimizuS, TomidaS, NiwaT, YokoyamaT, FukudaY, ChibaY, KatoR, TanizakiJ, TanakaK, TakedaM, OguraT, IshidaT, ItoA, and NakagawaK: Impact of EGFR-TKI treatment on the tumor immune microenvironment in EGFR mutation–positive non–small cell lung cancer. Clin Cancer Res, 2020; 26:2037–2046.
11.
ShiP, OhYT, DengL, ZhangG, QianG, ZhangS, RenH, WuG, LegendreBJr., AndersonE, RamalingamSS, OwonikokoTK, ChenM, and SunSY: Overcoming acquired resistance to AZD9291, a third-generation EGFR inhibitor, through modulation of MEK/ERK-dependent Bim and Mcl-1 degradation. Clin Cancer Res, 2017; 23:6567–6579.
12.
ShiP, OhYT, ZhangG, YaoW, YueP, LiY, KantetiR, RiehmJ, SalgiaR, OwonikokoTK, RamalingamSS, ChenM, and SunSY: Met gene amplification and protein hyperactivation is a mechanism of resistance to both first and third generation EGFR inhibitors in lung cancer treatment. Cancer Lett, 2016; 380:494–504.
13.
ZangH, QianG, ZongD, FanS, OwonikokoTK, RamalingamSS, and SunSY: Overcoming acquired resistance of epidermal growth factor receptor-mutant non-small cell lung cancer cells to osimertinib by combining osimertinib with the histone deacetylase inhibitor panobinostat (LBH589). Cancer, 2020; 126:2024–2033.
14.
ZhangT, QuR, ChanS, LaiM, TongL, FengF, ChenH, SongT, SongP, BaiG, LiuY, WangY, LiY, SuY, ShenY, SunY, ChenY, GengM, DingK, DingJ, and XieH: Discovery of a novel third-generation EGFR inhibitor and identification of a potential combination strategy to overcome resistance. Mol Cancer, 2020; 19:90.
15.
JonkerDJ, O'CallaghanCJ, KarapetisCS, ZalcbergJR, TuD, AuH-J, BerrySR, KrahnM, PriceT, SimesRJ, TebbuttNC, Van HazelG, WierzbickiR, LangerC, and MooreMJ: Cetuximab for the treatment of colorectal cancer. N Engl J Med, 2007; 357:2040–2048.
16.
WeberJ, and McCormackPL: Panitumumab: In metastatic colorectal cancer with wild-type KRAS. BioDrugs, 2008; 22:403–411.
17.
LiS, SchmitzKR, JeffreyPD, WiltziusJJ, KussieP, and FergusonKM: Structural basis for inhibition of the epidermal growth factor receptor by cetuximab. Cancer Cell, 2005; 7:301–311.
18.
ChenZ-P, YangQ, and GuoC-C: Profile of nimotuzumab in the treatment of high-grade glioma. Onco Targets Ther, 2015:8:819–825.
19.
HirschF, and GenovaC: Clinical potential of necitumumab in non-small cell lung carcinoma. Onco Targets Ther, 2016; 9:5427–5437.
20.
Crombet RamosT, Mestre FernándezB, Mazorra HerreraZ, and Iznaga EscobarNE: Nimotuzumab for patients with inoperable cancer of the head and neck. Front Oncol, 2020; 10:817.
21.
MaronSB, AlpertL, KwakHA, LomnickiS, ChaseL, XuD, O'DayE, NagyRJ, LanmanRB, CecchiF, HembroughT, SchrockA, HartJ, XiaoS-Y, SetiaN, and CatenacciDVT: Targeted therapies for targeted populations: Anti-EGFR treatment for EGFR-amplified gastroesophageal adenocarcinoma. Cancer Discov, 2018; 8:696–713.
22.
CaiWQ, ZengLS, WangLF, WangYY, ChengJT, ZhangY, HanZW, ZhouY, HuangSL, WangXW, PengXC, XiangY, MaZ, CuiSZ, and XinHW: The latest battles between EGFR monoclonal antibodies and resistant tumor cells. Front Oncol, 2020; 10:1249.
23.
BagchiA, HaidarJN, EastmanSW, ViethM, TopperM, IacolinaMD, WalkerJM, ForestA, ShenY, NovosiadlyRD, and FergusonKM: Molecular basis for necitumumab inhibition of EGFR variants associated with acquired cetuximab resistance. Mol Cancer Ther, 2018; 17:521–531.
24.
SickmierEA, KurzejaRJ, MichelsenK, VazirM, YangE, and TaskerAS: The panitumumab EGFR complex reveals a binding mechanism that overcomes cetuximab induced resistance. PLoS One, 2016; 11:e0163366.
25.
TalaveraA, FriemannR, Gomez-PuertaS, Martinez-FleitesC, GarridoG, RabasaA, Lopez-RequenaA, PupoA, JohansenRF, SanchezO, KrengelU, and MorenoE: Nimotuzumab, an antitumor antibody that targets the epidermal growth factor receptor, blocks ligand binding while permitting the active receptor conformation. Cancer Res, 2009; 69:5851–5859.
26.
VoigtM, BraigF, GöthelM, SchulteA, LamszusK, BokemeyerC, and BinderM: Functional dissection of the epidermal growth factor receptor epitopes targeted by panitumumab and cetuximab. Neoplasia, 2012; 14:1023–1031.
27.
ForsströmB, Bisławska AxnäsB, RockbergJ, DanielssonH, BohlinA, and UhlenM: Dissecting antibodies with regards to linear and conformational epitopes. PLoS One, 2015; 10:e0121673.
28.
Toride KingM, and BrooksCL.Epitope Mapping of Antibody-Antigen Interactions with X-Ray Crystallography. Springer, New York, 2018, pp. 13–27.
29.
LongF, FongRH, AustinSK, ChenZ, KloseT, FokineA, LiuY, PortaJ, SapparapuG, AkahataW, DoranzBJ, CroweJE, DiamondMS, and RossmannMG: Cryo-EM structures elucidate neutralizing mechanisms of anti-chikungunya human monoclonal antibodies with therapeutic activity. Proc Natl Acad Sci U S A, 2015; 112:13898–13903.
30.
BenjaminDC, and PerdueSS: Site-directed mutagenesis in epitope mapping. Methods, 1996; 9:508–515.
31.
DavidsonE, and DoranzBJ: A high-throughput shotgun mutagenesis approach to mapping B-cell antibody epitopes. Immunology, 2014; 143:13–20.
32.
WeiH, MoJ, TaoL, RussellRJ, TymiakAA, ChenG, IacobRE, and EngenJR: Hydrogen/deuterium exchange mass spectrometry for probing higher order structure of protein therapeutics: Methodology and applications. Drug Discov Today, 2014; 19:95–102.
33.
BourquardT, MusnierA, PuardV, TahirS, AyoubMA, JullianY, BouloT, GallayN, WatierH, BruneauG, ReiterE, CrépieuxP, and PouponA: MAbTope: A method for improved epitope mapping. J Immunol, 2018; 201:3096–3105.
34.
AsanoT, KanekoMK, and KatoY: RIEDL tag: A novel pentapeptide tagging system for transmembrane protein purification. Biochem Biophys Rep, 2020; 23:100780.
35.
ItaiS, YamadaS, KanekoMK, ChangYW, HaradaH, and KatoY: Establishment of EMab-134, a sensitive and specific anti-epidermal growth factor receptor monoclonal antibody for detecting squamous cell carcinoma cells of the oral cavity. Monoclon Antib Immunodiagn Immunother, 2017; 36:272–281.
36.
FujiiY, KanekoMK, and KatoY: MAP tag: A novel tagging system for protein purification and detection. Monoclon Antib Immunodiagn Immunother, 2016; 35:293–299.
37.
FujiiY, KanekoM, NeyazakiM, NogiT, KatoY, and TakagiJ: PA tag: A versatile protein tagging system using a super high affinity antibody against a dodecapeptide derived from human podoplanin. Protein Expr Purif, 2014; 95:240–247.
38.
KatoY, KanekoMK, KunoA, UchiyamaN, AmanoK, ChibaY, HasegawaY, HirabayashiJ, NarimatsuH, MishimaK, and OsawaM: Inhibition of tumor cell-induced platelet aggregation using a novel anti-podoplanin antibody reacting with its platelet-aggregation-stimulating domain. Biochem Biophys Res Commun, 2006; 349:1301–1307.
39.
KanekoMK, YamadaS, ItaiS, ChangYW, NakamuraT, YanakaM, and KatoY: Elucidation of the critical epitope of an anti-EGFR monoclonal antibody EMab-134. Biochem Biophys Rep, 2018; 14:54–57.
