Fractionation-Dependent Radiosensitization by Molecular Targeting of Nek1

Abstract

NIMA (never-in-mitosis gene A)-related kinase 1 (Nek1) is shown to impact on different cellular pathways such as DNA repair, checkpoint activation, and apoptosis. Its role as a molecular target for radiation sensitization of malignant cells, however, remains elusive. Stably transduced doxycycline (Dox)-inducible Nek1 shRNA HeLa cervix and siRNA-transfected HCT-15 colorectal carcinoma cells were irradiated in vitro and 3D clonogenic radiation survival, residual DNA damage, cell cycle distribution, and apoptosis were analyzed. Nek1 knockdown (KD) sensitized both cell lines to ionizing radiation following a single dose irradiation and more pronounced in combination with a 6 h fractionation (3 × 2 Gy) regime. For preclinical analyses we focused on cervical cancer. Nek1 shRNA HeLa cells were grafted into NOD/SCID/IL-2Rγc−/− (NSG) mice and Nek1 KD was induced by Dox-infused drinking water resulting in a significant cytostatic effect if combined with a 6 h fractionation (3 × 2 Gy) regime. In addition, we correlated Nek1 expression in biopsies of patients with cervical cancer with histopathological parameters and clinical follow-up. Our results indicate that elevated levels of Nek1 were associated with an increased rate of local or distant failure, as well as with impaired cancer-specific and overall survival in univariate analyses and for most endpoints in multivariable analyses. Finally, findings from The Cancer Genome Atlas (TCGA) validation cohort confirmed a significant association of high Nek1 expression with a reduced disease-free survival. In conclusion, we consider Nek1 to represent a novel biomarker and potential therapeutic target for drug development in the context of optimized fractionation intervals.

NEK1 Phosphorylation of YAP Promotes Its Stabilization and Transcriptional Output

“…Findings from our lab and others suggest that ADT activates the TLK1-NEK1 signaling pathway that promotes PCa progression by activating the DDR [11,24]. Hyper-activation of NEK1 may also lengthen G2/M checkpoints, which provides the cells sufficient time to repair their damaged DNA after ADT or radiation therapy [7,58]. However, DDR alone may not be able to induce androgen-insensitive growth of PCa cells.…”

Analysis of the effect of NEKs on the prognosis of patients with non-small-cell lung carcinoma based on bioinformatics

“…When it is mutated or silenced, spindle defects, abnormal chromosome segregation, mitotic block, and apoptosis occur; therefore, NEK1 plays an important role in the development of cancer 34 , 58 – 60 . NEK1 can participate in the regulation of DNA damage repair in HeLa cervical cancer cells 61 , and its overexpression can significantly reduce disease-free survival 62 ; it can also be overexpressed in human glioma tissues, which is related to tumor grade and nodal metastasis status 63 . A decrease in NEK10 expression can lead to increased cell proliferation and DNA replication 64 , which is related to poor prognosis and higher tumor grade of breast cancer 65 .…”

Illuminating a Dark Kinase: Structure-Guided Design, Synthesis, and Evaluation of a Potent Nek1 Inhibitor and Its Effects on the Embryonic Zebrafish Pronephros

“…Furthermore, the knockdown sensitized those cells to ionizing radiation, underlining the potential of Nek1 inhibitors to increase the radiation or chemotherapy response in anticancer treatment. 30 Despite the crucial role of Nek1 in a broad variety of diseases and its outstanding potential as a target for anticancer therapeutics, no directed medicinal chemistry efforts toward potent and selective Nek1 inhibitors have been published in the past. 4,31 In a first attempt to pinpoint structural elements that influence Nek family inhibitor specificity, Moraes et al identified a c-Jun N-terminal kinase (JNK) inhibitor with limited inhibitory activity against a truncated Nek1 activation loop mutant in a thermal shift assay.…”

“…Direct phosphorylation of voltage-dependent anion channel 1 (VDAC1) by Nek1 stabilizes the mitochondrial membrane potential thus limiting the caspase-mediated killing of cells. , More recent evidence suggests that an activating phosphorylation of Nek1 by tousled-like kinase 1 (Tlk1) contributes to the phosphorylation of VDAC1 and that inhibition of the Tlk1/Nek1/VDAC1 axis in combination with DNA damaging agents can sensitize prostate cancer cells to apoptotic killing. − Elevated levels of Nek1 expression have also been identified in other cancer types like thyroid cancer, human gliomas, or renal cell carcinoma (RCC). − In the latter case, a decreased sensitivity to genotoxic treatment and ionizing radiation was associated with high levels of Nek1, emphasizing the importance of Nek1 inhibition as a therapeutic strategy for drug development in the treatment of RCC. , More recently, Freund et al have shown that high Nek1 expression levels associate with impaired clinical outcome in cervical cancer patients following chemo-radiotherapy or brachytherapy and that a Nek1 knockdown impacts the 3D clonogenic survival of HeLa cervical and HCT-15 colorectal carcinoma cells. Furthermore, the knockdown sensitized those cells to ionizing radiation, underlining the potential of Nek1 inhibitors to increase the radiation or chemotherapy response in anticancer treatment …”