Shao Hanshuang, Wells Alan
Department of Pathology, University of Pittsburgh, Pittsburgh, PA 15213, USA.
Pittsburgh VA Health System, Pittsburgh, PA 15213, USA.
Int J Mol Sci. 2025 Jun 26;26(13):6157. doi: 10.3390/ijms26136157.
Abnormal expressions and genetic mutations of EGFR are broadly involved in the progression of many human solid tumors, which has led to the development of small molecule inhibitors (TKIs). However, patients' tumors usually develop resistance to targeted therapeutic TKIs after a period of treatment, mostly due to secondary mutations in EGFR. To date, three major and prevalent point mutations in EGFR, including L858R, T790M, and C797S, impact the use of TKIs in non-small cell lung cancer patients. Although at least four generations of TKIs have been designed and developed by targeting these mutations, how each mono, dual, or triple variant responds to clinical TKIs remains largely undeciphered. To fill this gap, we constructed a series of EGFR mutants and assessed their responses to clinical TKIs in vitro. The first-generation TKI, erlotinib, completely blocked the autophosphorylation of WT, L858R, C797S, and C797S/L858R, but only partially, if at all, in EGFR containing the T790M mutation alone or in combination. The third generation, osimertinib, completely abolished the autophosphorylation of WT, T790M, L858R, and T790M/L858R. It also significantly inhibited C797S and C790S/L858R, but had no effect on T790M/C797S or T790M/C797S/L858R. EAI045, as the fourth-generation TKI, almost completely inhibited WT and all mutants in complete growth media, but EGF-mediated phosphorylation of WT, C797S, and C797S/L858R were only partially inhibited in quiescence media, while the other mutants were fully inhibited. Furthermore, the abolishment of the enhanced tolerance to Dox in cells transiently expressing T790M/L858R and T790M/C797S/L858R by EAI045 suggests that their enhanced autophosphorylation is involved in their resistant ability. These findings provide some insights into how patients carrying typical mutations should be correctly and efficiently treated and why patients present side effects (because of non-specific inhibitory effects on cells without EGFR mutations).
表皮生长因子受体（EGFR）的异常表达和基因突变广泛参与多种人类实体瘤的进展，这促使了小分子抑制剂（TKIs）的研发。然而，患者的肿瘤在经过一段时间的治疗后通常会对靶向治疗性TKIs产生耐药性，这主要是由于EGFR的二次突变所致。迄今为止，EGFR中的三种主要且常见的点突变，包括L858R、T790M和C797S，影响了TKIs在非小细胞肺癌患者中的应用。尽管针对这些突变已经设计并开发了至少四代TKIs，但每种单变体、双变体或三变体对临床TKIs的反应在很大程度上仍未得到破解。为了填补这一空白，我们构建了一系列EGFR突变体，并在体外评估了它们对临床TKIs的反应。第一代TKI厄洛替尼完全阻断了野生型、L858R、C797S和C797S/L858R的自磷酸化，但对于仅含有T790M突变或与其他突变组合的EGFR，其阻断作用仅为部分，甚至完全没有作用。第三代奥希替尼完全消除了野生型、T790M、L858R和T790M/L858R的自磷酸化。它还显著抑制了C797S和C790S/L858R，但对T790M/C797S或T790M/C797S/L858R没有作用。作为第四代TKI的EAI045在完全生长培养基中几乎完全抑制了野生型和所有突变体，但在静止培养基中，EAI045仅部分抑制了野生型、C797S和C797S/L858R的表皮生长因子（EGF）介导的磷酸化，而对其他突变体则完全抑制。此外，EAI045消除了瞬时表达T790M/L858R和T790M/C797S/L858R的细胞对阿霉素增强的耐受性，这表明它们增强的自磷酸化与耐药能力有关。这些发现为如何正确、有效地治疗携带典型突变的患者以及患者为何会出现副作用（由于对没有EGFR突变的细胞产生非特异性抑制作用）提供了一些见解。
