Yu Yang, Zhao Yanling, Shi Zhongcheng, Cheng Feng, Wang Larry L, Choi Jong Min, Li Kan, Silverman Daniel, Qi Dan, Wang Jun, Agarwal Saurabh, Rood Brian R, Dome Jeffrey S, Fabbri Muller, Yi Joanna S, Wu Erxi, Jung Sung Yun, Zhang Chunchao, Yang Jianhua
Center for Cancer and Immunology Research, Children's National Research Institute, Children's National Hospital, Washington, DC 20010, USA.
Texas Children's Hospital, Department of Pediatrics, Dan L. Duncan Cancer Center, Baylor College of Medicine, Houston, TX 77030, USA.
bioRxiv. 2024 Feb 22:2024.02.19.581040. doi: 10.1101/2024.02.19.581040.
Neuroblastoma (NB) can be a highly aggressive malignancy in children. However, the precise mechanisms driving NB tumorigenesis remain elusive. This study revealed the critical role of CREB phosphorylation in NB cell proliferation. By employing a CRISPR-Cas9 knockout screen targeting calcium/calmodulin-dependent protein kinase (CaMK) family members, we identified the CaM kinase-like vesicle-associated (CAMKV) protein as a kinase that mediates direct phosphorylation of CREB to promote NB cell proliferation. was found to be a transcriptional target of MYCN/MYC in NB cells. CAMKV knockout and knockdown effectively suppressed NB cell proliferation and tumor growth both in vitro and in vivo. Bioinformatic analysis revealed that high CAMKV expression is significantly correlated with poor patient survival. High-risk NB frequently had high CAMKV protein levels by Immunohistochemical staining. Integrated transcriptomic and proteomic analyses of CAMKV knockdown cells unveiled downstream targets involved in CAMKV-regulated phosphorylation and signaling pathways, many of which are linked to neural development and cancer progression. We identified small molecule inhibitors targeting CAMKV and further demonstrated the efficacy of one inhibitor in suppressing NB tumor growth and prolonging the survival of mice bearing xenografted tumors. These findings reveal a critical role for CAMKV kinase signaling in NB growth and identified CAMKV kinase as a potential therapeutic target and prognostic marker for patients with NB.
神经母细胞瘤（NB）在儿童中可能是一种高度侵袭性的恶性肿瘤。然而，驱动NB肿瘤发生的确切机制仍不清楚。本研究揭示了CREB磷酸化在NB细胞增殖中的关键作用。通过使用针对钙/钙调蛋白依赖性蛋白激酶（CaMK）家族成员的CRISPR-Cas9基因敲除筛选，我们确定钙调素激酶样囊泡相关蛋白（CAMKV）作为一种激酶，介导CREB的直接磷酸化以促进NB细胞增殖。发现CAMKV是NB细胞中MYCN/MYC的转录靶点。CAMKV基因敲除和敲低在体外和体内均有效抑制NB细胞增殖和肿瘤生长。生物信息学分析显示，高CAMKV表达与患者生存率低显著相关。通过免疫组织化学染色发现，高危NB经常具有高CAMKV蛋白水平。对CAMKV敲低细胞的综合转录组学和蛋白质组学分析揭示了参与CAMKV调节的磷酸化和信号通路的下游靶点，其中许多靶点与神经发育和癌症进展有关。我们鉴定了靶向CAMKV的小分子抑制剂，并进一步证明了一种抑制剂在抑制NB肿瘤生长和延长荷瘤小鼠生存期方面的功效。这些发现揭示了CAMKV激酶信号在NB生长中的关键作用，并确定CAMKV激酶是NB患者潜在的治疗靶点和预后标志物。
