Exportin 1 inhibition prevents neuroendocrine transformation through SOX2 downregulation in lung and prostate cancers [RNA-seq]

In lung and prostate adenocarcinomas, neuroendocrine (NE) transformation to an aggressive derivative resembling small cell lung cancer (SCLC) is associated with poor prognosis. We previously described dependency of SCLC on the nuclear transporter exportin 1. Here we explored the role of exportin 1 in NE transformation. We observed upregulated exportin 1 in lung and prostate pre-transformation adenocarcinomas. Exportin 1 was induced upregulated following genetic inactivation of TP53 and RB1 in lung and prostate adenocarcinoma cell lines, accompanied by increased sensitivity to the exportin 1 inhibitor selinexor in vitro. Exportin 1 inhibition prevented NE transformation and extended response to targeted therapies in both lung anddifferent TP53/RB1-inactivated prostate adenocarcinoma xenograft models that acquire NE features upon treatment with the AR inhibitor enzalutamide, and extended response to the EGFR inhibitor osimertinib in a lung cancer transformation patient-derived xenograft (PDX) model exhibiting combined adenocarcinoma/SCLC histology. Ectopic SOX2 expression restored the enzalutamide-promoted NE transformationNE phenotype on adenocarcinoma-to-NE transformation xenograft models despite selinexor treatment. Selinexor sensitized NE-transformed lung and prostate small cell carcinoma PDXs tumors after NE transformation to standard cytotoxics. Together these data nominate exportin 1 inhibition as a novel potential therapeutic approach target to constrain lineage plasticity and prevent or treat NE transformation in lung and prostate adenocarcinoma. 4-10 female 6-week-old NOD.Cg-Prkdc Il2rg/SzJ (NSG) mice were subcutaneously engrafted per treatment arm and until tumors reached 100-150 mm3. At that point, mice were randomized into groups and treated with either vehicle, cisplatin (2 mg/kg intraperitoneally (i.p.) once/week), etoposide (3 mg/kg i.p. QDx3), selinexor (10 mg/kg p.o. QDx3), enzalutamide (10 mg/kg per os (p.o). QDx5), osimertinib (25 mg/kg p.o. QDx5) or the combinations of cisplatin + etoposide, cisplatin + selinexor, enzalutamide + selinexor or osimertinib + selinexor at the previously mentioned doses. 3-4 tumors per treatment group were collected at day 56 (day 31 for the control condition). Mice weights and tumor volumes were measured twice a week and mice were sacrificed when tumors reached humane endpoint (volume = 1000 mm3). The number of mice per treatment arm were selected according to previous experience with the models and response to treatments. Blinding was not performed. All animal experiments were approved by the Memorial Sloan Kettering Cancer Center (MSKCC) Animal Care and Use Committee (#13-07-007). Frozen tissues were weighed and homogenized in RLT and nucleic acids were extracted using the AllPrep DNA/RNA Mini Kit (QIAGEN, #80204) according to the manufacturer’s instructions. RNA was eluted in nuclease-free water. **Raw data not provided due to accidental deletion of files**在肺腺癌与前列腺腺癌中，向类似小细胞肺癌（small cell lung cancer, SCLC）的侵袭性表型发生的神经内分泌（neuroendocrine, NE）转化，与不良预后密切相关。我们此前已阐明小细胞肺癌对核转运蛋白exportin 1的依赖性。本研究探讨了exportin 1在神经内分泌转化中的作用。我们在肺与前列腺的癌前腺癌中观察到exportin 1表达上调。在肺腺癌细胞系与前列腺腺癌细胞系中，TP53与RB1基因失活后可诱导exportin 1表达上调，同时使细胞对exportin 1抑制剂selinexor的体外敏感性升高。在肺腺癌和经雄激素受体（AR）抑制剂恩扎卢胺（enzalutamide）处理后获得神经内分泌特征的不同TP53/RB1失活型前列腺腺癌异种移植模型中，exportin 1抑制可阻断神经内分泌转化，并延长对靶向治疗的应答；在表现出腺癌/SCLC混合组织学特征的肺癌转化患者来源异种移植（patient-derived xenograft, PDX）模型中，exportin 1抑制可延长对EGFR抑制剂奥希替尼（osimertinib）的应答。尽管接受selinexor治疗，异位过表达SOX2仍可在腺癌向神经内分泌转化的异种移植模型中恢复恩扎卢胺诱导的神经内分泌转化表型。Selinexor可使经神经内分泌转化后的肺与前列腺小细胞癌PDX肿瘤对标准细胞毒性化疗敏感。综上，本研究数据将exportin 1抑制列为一种潜在的新型治疗策略，旨在限制谱系可塑性，预防或治疗肺腺癌与前列腺腺癌中的神经内分泌转化。 每治疗组皮下接种4~10只6周龄雌性NOD.Cg-Prkdc Il2rg/SzJ（NSG）小鼠，直至肿瘤体积达到100~150 mm³。此时将小鼠随机分组，分别给予赋形剂、顺铂（2 mg/kg，腹腔注射，每周1次）、依托泊苷（3 mg/kg，腹腔注射，每3天1次，共3次）、selinexor（10 mg/kg，口服，每3天1次，共3次）、恩扎卢胺（10 mg/kg，口服，每5天1次，共5次）、奥希替尼（25 mg/kg，口服，每5天1次，共5次），或顺铂+依托泊苷、顺铂+selinexor、恩扎卢胺+selinexor、奥希替尼+selinexor联合治疗，给药剂量同上。每治疗组在第56天收集3~4个肿瘤（对照组于第31天收集）。每周测量两次小鼠体重与肿瘤体积，当肿瘤达到人道终点（体积=1000 mm³）时处死小鼠。每组小鼠数量根据既往该模型的实验经验及治疗应答情况确定。本实验未设盲法。所有动物实验均经纪念斯隆凯特琳癌症中心（Memorial Sloan Kettering Cancer Center, MSKCC）动物护理与使用委员会批准（编号#13-07-007）。 冷冻组织称重后于RLT缓冲液中匀浆，按照制造商说明书使用AllPrep DNA/RNA Mini Kit（QIAGEN，#80204）提取核酸。RNA经无核酸酶水洗脱。**原始数据因文件意外删除未提供**