Discovery of Spirosnuolides A–D, Type I/III Hybrid Polyketide Spiro-Macrolides for a Chemotherapeutic Lead against Lung Cancer

Four new macrolides, spirosnuolides A–D (1–4, respectively), were discovered from the termite nest-derived Kitasatospora sp. INHA29. Spirosnuolides A–D are 18-membered macrolides sharing an embedded [6,6]-spiroketal functionality inside the macrocycle and are conjugated with structurally uncommon side chains featuring cyclopentenone, 1,4-benzoquinone, hydroxyfuroic acid, or butenolide moieties. Structure elucidation was achieved using a combination of spectroscopic analyses, multiple chemical derivatizations (methylation, methanolysis, Luche reduction, and Mosher’s reaction), X-ray diffraction analysis, and computational ECD calculations. Interestingly, genome sequencing analysis suggests that spirosnuolides were biosynthesized through a rare type I/III hybrid polyketide synthase. Importantly, spirosnuolide B displayed potent antiproliferative effects against various cancer cell lines at nanomolar concentrations, particularly against HCC827 cells, an EGFR mutant non-small-cell lung cancer (NSCLC) cell line, with a high safety index value. Based on in vitro studies, the antiproliferative mechanism of spirosnuolide B involved the activation of AMPK signaling, leading to cell cycle arrest and apoptosis in HCC827 cells. Its potent efficacy was also proven in vivo by the effective inhibition of tumor growth in mouse xenograft studies. Moreover, cotreatment with spirosnuolide B and gefitinib, synergistically enhanced the antiproliferative activity and apoptosis, suggesting a potential strategy to overcome gefitinib resistance in EGFR mutant NSCLC.

本研究从白蚁巢来源的北里孢菌属（Kitasatospora）菌株INHA29中，分离得到4种全新的大环内酯类（macrolides）化合物——螺螺诺内酯（spirosnuolides）A-D（分别对应化合物1~4）。螺螺诺内酯A-D均为18元大环内酯类化合物，其大环骨架内嵌有[6,6]-螺环缩酮官能团，并连有带有环戊烯酮、1,4-苯醌、羟基呋喃甲酸或丁烯内酯基团的罕见侧链。本研究通过光谱分析、多种化学衍生化手段（甲基化、甲醇解、Luche还原及Mosher’s反应）、X射线衍射分析以及电子圆二色谱（electronic circular dichroism, ECD）计算相结合的方法，完成了所有化合物的结构解析。有趣的是，基因组测序分析结果显示，螺螺诺内酯的生物合成依赖于一种罕见的I/III型杂合聚酮合酶（polyketide synthase, PKS）。值得关注的是，螺螺诺内酯B在纳摩尔浓度下即可对多种癌细胞系展现出强效抗增殖活性，尤其对表皮生长因子受体（epidermal growth factor receptor, EGFR）突变型非小细胞肺癌（non-small-cell lung cancer, NSCLC）细胞系HCC827活性突出，且具有较高的安全指数。体外实验表明，螺螺诺内酯B的抗增殖机制通过激活腺苷酸活化蛋白激酶（adenosine monophosphate-activated protein kinase, AMPK）信号通路，诱导HCC827细胞发生细胞周期阻滞与细胞凋亡。小鼠异种移植模型实验进一步证实了其体内抗肿瘤活性，可有效抑制肿瘤生长。此外，螺螺诺内酯B与吉非替尼联合给药可协同增强抗增殖活性并诱导细胞凋亡，为克服EGFR突变型非小细胞肺癌的吉非替尼耐药性提供了潜在治疗策略。