克服先天/后天药物耐药性的缺乏抗癌药物是实现癌症治疗持久完全缓解的最大障碍。为了解决这个问题，我们开发了一种新的药物系列，将生物活性氨基硫醇WR1065与离散的巯基-聚乙二醇聚合物结合起来进行细胞内传递：4星-聚乙二醇-二硫-WR1065（4SP65）每个分子释放四个WR1065，m-聚乙二醇6-二硫-WR1065（1LP65）每个分子释放一个WR1065。有时，通过经过FDA批准的细胞保护剂阿米法汀以细胞外方式提供一个WR1065分子，WR1065显示出抗癌效果。在由七种组织衍生的15种人类癌细胞系panel中评估了4SP65、1LP65和阿米法汀的相对抗癌效果。其他实验评估了4SP65联合治疗增强抗癌效果、克服顺铂（一种化疗药物）或吉非替尼（针对癌基因EGFR突变的酪氨酸激酶抑制剂）的耐药性的能力。使用CyQUANT®-NF增殖实验评估了48小时药物处理后的细胞存活率，并使用国家癌症研究所的COMPARE方法评估剂量-反应关系。在正常的人类上皮细胞中，4SP65或1LP65能够促进或抑制细胞生长，但不具有细胞毒性。在癌细胞系中，4SP65和1LP65能够诱导剂量依赖的细胞阻滞和溶解作用，在药物浓度为11.2±1.2 µM和126±15.8 µM时，分别达到了99%的细胞死亡。阿米法汀对11/14种癌细胞系的细胞阻滞作用有限，没有细胞溶解作用。4SP65加顺铂或吉非替尼的二元组合增强了每种合作药物的效果，并克服了相关癌细胞系对顺铂和吉非替尼细胞溶解的耐药性。4SP65和1LP65对TP53突变细胞系比对TP53野生型细胞系具有显著更高的效果，与WR1065介导的突变p53重激活一致。因此，4SP65和1LP65代表了一种独特的前药家族，可作为广谱抗癌药物应用在创新领域，目标是p53，并与化疗药物和EGFR-TKI协同作用，以预防或克服药物耐药性。 版权所有 © 2023 Walker、Lazarova、Riesinger、Poirier、Messier、Cunniff和Walker。

The lack of anticancer agents that overcome innate/acquired drug resistance is the single biggest barrier to achieving a durable complete response to cancer therapy. To address this issue, a new drug family was developed for intracellular delivery of the bioactive aminothiol WR1065 by conjugating it to discrete thiol-PEG polymers: 4-star-PEG-S-S-WR1065 (4SP65) delivers four WR1065s/molecule and m-PEG6-S-S-WR1065 (1LP65) delivers one. Infrequently, WR1065 has exhibited anticancer effects when delivered via the FDA-approved cytoprotectant amifostine, which provides one WR1065/molecule extracellularly. The relative anticancer effectiveness of 4SP65, 1LP65, and amifostine was evaluated in a panel of 15 human cancer cell lines derived from seven tissues. Additional experiments assessed the capacity of 4SP65 co-treatments to potentiate the anticancer effectiveness and overcome drug resistance to cisplatin, a chemotherapeutic, or gefitinib, a tyrosine kinase inhibitor (TKI) targeting oncogenic EGFR mutations. The CyQUANT®-NF proliferation assay was used to assess cell viability after 48-h drug treatments, with the National Cancer Institute COMPARE methodology employed to characterize dose-response metrics. In normal human epithelial cells, 4SP65 or 1LP65 enhanced or inhibited cell growth but was not cytotoxic. In cancer cell lines, 4SP65 and 1LP65 induced dose-dependent cytostasis and cytolysis achieving 99% cell death at drug concentrations of 11.2 ± 1.2 µM and 126 ± 15.8 µM, respectively. Amifostine had limited cytostatic effects in 11/14 cancer cell lines and no cytolytic effects. Binary pairs of 4SP65 plus cisplatin or gefitinib increased the efficacy of each partner drug and surmounted resistance to cytolysis by cisplatin and gefitinib in relevant cancer cell lines. 4SP65 and 1LP65 were significantly more effective against TP53-mutant than TP53-wild-type cell lines, consistent with WR1065-mediated reactivation of mutant p53. Thus, 4SP65 and 1LP65 represent a unique prodrug family for innovative applications as broad-spectrum anticancer agents that target p53 and synergize with a chemotherapeutic and an EGFR-TKI to prevent or overcome drug resistance.Copyright © 2023 Walker, Lazarova, Riesinger, Poirier, Messier, Cunniff and Walker.