The expression of the erythroblastic leukemia viral oncogene homolog (ERBB) family is closely linked to tumor progression through the constitutive activation of downstream signaling, such as the epidermal growth factor (EGF) receptor (EGFR) pathway or through a somatic mutation; ERBB expression is enhanced during tumor microenvironment (TME) formation, cancer progression, and drug-resistance
[1]. The ERBB family comprises transmembrane receptor tyrosine kinases, including the ERBB1/ EGFR)/HER (human EGF receptor) 1, ERBB2/HER2/Neu, ERBB3/HER3, and ERBB4/HER4
[2]. Activated ERBB mediates various signaling pathways including the RAS (rat sarcoma)/RAF (rapidly accelerated fibrosarcoma), phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K)/AKT (a serine/threonine protein kinase), phospholipase C (PLC)-γ1, and signal transducer and activator of transcription (STAT) pathways
[3]. Several ligands can bind to EGFR, including EGF, epigen, transforming growth factor (TGF)-α, amphiregulin (AREG), epiregulin (EREG), betacellulin (BTC), and heparin-binding EGF (HB-EGF) (
Figure 1). The simplified representation of these ligands with distinct functional domains are shown in
Figure 1. The transmembrane EGFR ligands comprise an N-terminal signal peptide, pro-peptide region, the EGF-lik short juxtamembrane stalk, a hydrophobic transmembrane domain, and a cytoplasmic domain (
Figure 1A). Neuregulins (NRGs) are a family containing the EGF-like domain proteins; they play an essential role in the development of the adult brain
[4] and various cancers
[5][6][7][5,6,7]. The most studied NRGs, such as the
NRG1 gene, produce six different types and 33 spliced isoforms, due to different transcriptional initiation sites and alternative splicing
[4]. NRG proteins mainly contain EGF-like and transmembrane domains; however, the type-specific N-terminal region (type I, II, and IV-VI NRG1), an immunoglobulin-like domain, and the glycosylation site are dependent on the isoform
[8][9][8,9]. Additionally, the identity of the overall protein sequence in these ligands is low
[10] and a conserved EGF module including six cysteines is arranged as three disulfide bridges (
Figure 1B). The spacing of EGF-motif in seven EGFR ligands can be represented as in the cysteines pattern CX
7CX
4–5CX
10CXCX
8C (X can be any amino acid). Notably, recent studies demonstrated that the N57 residue of EREG is pivotal for the interaction with the domains I and III of EGFR
[11]. EGF is unique in that there are nine EGF motifs, although only the one adjacent to the cell membrane has the function of the EGFR binding domain (
Figure 1C). EREG and HB-EGF contain an additional heparin-binding domain. The functional EGF module is located within approximately 25 residues of the transmembrane domain. The presence and spacing of additional specific residues further distinguishes EGFR ligands from NRGs containing EGF modules at the structural level, and defines high-affinity binding to EGFR
[12]. Moreover, NRG1 and 2 selectively bind to ERBB3 (
Figure 1D). Ligands such as BTC, HB-EGF, EREG, and NRG1-4 interact with the ERBB4. ERBB ligands bind to the extracellular domain of ERBB1, ERBB3, and ERBB4 receptors to form active homodimers or heterodimers. However, ligands do not directly bind to ERBB2 in the ligand-activated state, favoring homodimerization. In addition, ERBB2 proteins can be activated through interaction with other ERBBs. G-protein-coupled receptors (GPCRs) stimulate specific metalloproteinases, such as disintegrin and metalloproteinase (ADAM) family members, resulting in EGFR pro-ligand cleavage and transactive EGFR downstream cascade
[13] (
Figure 1E). Ectodomain shedding arises in diverse physiological responses, and the cleavage efficiency is mainly determined by the specific sequence in the cleavage site and the length of the membrane-proximal domain
[14]. Soluble ligands bind to receptors, activating intracellular signaling on the original cell, neighboring cells, and distant cells through autocrine, paracrine (or juxtacrine), and endocrine pathways, respectively
[12]. EGFR-medaited signaling pathways can be activated by binding to soluble ligands or membrane-anchored ligands by the juxtacrine pathway. In addition to the actions of soluble ligands, the free cytoplasmic tail (CT) of these ligands (e.g., pro-AREG CT) are required for basolateral sorting
[15] and pro-HBE-GF CT can directly regulate gene expression
[16]. The efficacy and specificity of intracellular signaling pathways are regulated by specific ligands, receptor dimerization, and interacting proteins that bind to the phosphorylated domains of ERBB
[17].