Project 4: SIK2 PROVIDES A NOVEL TARGET FOR OVARIAN CANCER THERAPY IN COMBINATION WITH PACLITAXEL AND INHIBITORS OF PARP

项目4摘要/摘要 在过去的30年里，上皮性卵巢癌患者的5年生存率有所提高，但长期 长期存活率没有变化，总体上保持在30%左右。在手术细胞减少后，所有 新患者接受标准的初级化疗，包括卡铂和紫杉醇的组合。 仅用含铂化合物的初级治疗就能使大约70%的卵巢退化 而42%的人只对紫杉醇有反应，两种药物之间没有协同作用。我们的长- 术语翻译的目标是提高卵巢癌化疗的有效性。虽然很多人 研究人员一直专注于克服对紫杉烷类药物的获得性耐药性，但相对较少的关注 在初治化疗期间致力于增强紫杉醇的反应。使用功能性siRNA筛查 调节紫杉醇敏感性的激酶，我们发现丝氨酸-苏氨酸激酶盐的敲除- 诱导蛋白2(SIK2)诱导多倍体、抑制卵巢癌生长和增强紫杉醇敏感性 在细胞系和异种移植中。SIK2是正常中心体分裂所必需的，在34%的 所有组织类型的卵巢癌，与总存活率降低有关。在有丝分裂过程中，SIK2经历 自动磷酸化和磷酸化C-NAP1，这是一种介导中心体分裂的结构蛋白。SIK2 也使P85DNA磷酸化，驱动PI3K和α的激活，改变染色质和DNA修复。 我们已经与Arrien制药公司建立了合作关系，开发口服小剂量 单一溶剂结合不同的SIK2：ARN-3236和ARN-3261的分子量抑制剂 换人。ARN-3236对10种卵巢癌细胞株的IC50值为0.8~2.6ICM，其中μ-3236对10种卵巢癌细胞株的生长抑制作用最强。 ARN3236的IC50值与内源性SIK2的表达呈负相关(Pearson‘s r=-0.642，P=0.03)。 在10个细胞株中，有8个细胞株和SKOv3ip细胞对紫杉醇的敏感性增强(P=0.028)。 OVCAR8异种移植。ARN-3236使中心体在间期从细胞核解偶联，被阻断 有丝分裂中的中心体分离，导致前中期停滞，诱导细胞凋亡以及 四倍体。ARN-3236还可抑制AKT的磷酸化和抑制Survivin的表达。ARN-3261 在异种移植模型中增加了对紫杉醇和顺铂的敏感性，并且显示出很小的毒性以及 对PGP的抵抗力更强，效力是ARN-3236的13倍。我们将追求三个目标：1)履行 SIK2抑制剂ARN-3261单独(IA)和每周紫杉醇(IB)联合应用的I期试验 测定ARN-3261和紫杉醇的药代动力学、预测性(SIK2)和药效学(pSIK2和 PHDAC5)生物标志物，以及多倍体、PAKT、Survivin和凋亡生物标志物的水平；2)至 确定SIK2抑制剂ARN-3261是否增强对卡铂/紫杉醇和 卡铂/脂质体阿霉素在卵巢癌细胞系来源和患者来源的异种移植中的作用； 确定在用SIK2抑制剂治疗的卵巢癌细胞中产生合成杀伤力的靶点。

Project 4 SUMMARY/ABSTRACT Over the last three decades, 5-year survival has improved for patients with epithelial ovarian cancer, but long- term survival has not changed and remains at approximately 30% overall. Following surgical cytoreduction, all new patients receive standard primary chemotherapy that includes a combination of carboplatin and paclitaxel. Primary therapy with platinum-based compounds alone produces regression in approximately 70% of ovarian cancers, whereas 42% respond to paclitaxel alone and there is no synergy between the two drugs. Our long- term translational goal is to increase the effectiveness of chemotherapy for ovarian cancer. While many investigators have focused on overcoming acquired resistance to taxanes, relatively little attention has been given to enhancing paclitaxel response during primary chemotherapy. Using a functional siRNA screen for kinases that regulate sensitivity to paclitaxel, we found that knockdown of the serine-threonine kinase salt- induced kinase 2 (SIK2) induces polyploidy, inhibits ovarian cancer growth and enhances paclitaxel sensitivity in cell lines and xenografts. SIK2 is required for normal centrosome splitting and is overexpressed in 34% of ovarian cancers of all histotypes, associated with decreased overall survival. During mitosis, SIK2 undergoes autophosphorylation and phosphorylates C-Nap1 a structural protein that mediates centrosome splitting. SIK2 also phosphorylates p85α, driving activation of PI3K, as well as HDAC5, modifying chromatin and DNA repair. We have established a collaboration with Arrien Pharmaceuticals to develop orally administered small molecular weight inhibitors of SIK2: ARN-3236 and ARN-3261 that differ by a single solvent binding substitution. ARN-3236 inhibited growth of 10 ovarian cancer cell lines at an IC50 of 0.8 to 2.6 μM, where the IC50 of ARN-3236 was inversely correlated with endogenous SIK2 expression (Pearson’s r = -0.642, P = 0.03). ARN-3236 also enhanced sensitivity to paclitaxel in 8 of 10 cell lines, as well as in SKOv3ip (P = 0.028) and OVCAR8 xenografts. ARN-3236 uncoupled the centrosome from the nucleus in interphase, blocked centrosome separation in mitosis, caused prometaphase arrest and induced apoptotic cell death as well as tetraploidy. ARN-3236 also inhibited AKT phosphorylation and attenuated survivin expression. ARN-3261 enhanced sensitivity to paclitaxel and cisplatin in xenograft models and exhibited little toxicity, as well as greater resistance to PgP and13-fold greater potency than ARN-3236. We will pursue three aims: 1) to perform a phase I trial of the SIK2 inhibitor ARN-3261 alone (IA) and in combination with weekly paclitaxel (IB) measuring pharmacokinetics of ARN-3261 and paclitaxel, predictive (SIK2) and pharmacodynamic (pSIK2 and pHDAC5) biomarkers, as well as levels of polyploidy, pAKT, survivin and biomarkers for apoptosis; 2) to determine whether the SIK2 inhibitor ARN-3261 enhances response to carboplatin/paclitaxel and carboplatin/liposomal doxorubicin in ovarian cancer cell line-derived and patient-derived xenografts; and 3) to identify targets that produce synthetic lethality in ovarian cancer cells treated with SIK2 inhibitors.