CK1α-targeting inhibits primary and metastatic colorectal cancer in vitro, ex vivo, in cell-line-derived and patient-derived tumor xenograft mice models

Abstract

Background

Colorectal cancer (CRC) remains a leading cause of cancer-related deaths globally. Despite improved understanding of its initiation and progression, and advances in diagnostic or therapeutic strategies, the treatment of metastatic CRC remains a clinical challenge, necessitating identification of novel efficacious therapeutics with little/no toxicity to non-tumor colorectal cells. The present study investigated the effect of Epiblastin A, an adenosine triphosphate (ATP)-mediated competitive inhibitor of casein kinase 1α (CK1α) on the viability, proliferation, and oncogenicity of CRC cells.

Methods

Comparative evaluation of the effect of Epiblastin A on CK1α in fetal human normal colonic mucosa (FHC) and CRC (HCT116, HT29, DLD1) cell lines, using western blot, immunohistochemical staining, real-time polymerase chain reaction (RT-PCR), and sulforhodamine B (SRB) cytotoxicity assays. Primary culture cells, patient-derived xenograft (PDX), and tumor xenograft mice CRC models were also employed. Kaplan-Meier plots were used for survival analysis of our CRC cohort.

Results

CRC cells aberrantly express CK1α at mRNA and protein levels. This overexpression of CK1α is strongly associated with worse 5-year overall survival (OS) in patients with CRC. Epiblastin A inhibits CK1α and compared to its apparent non-effect on FHC cells regardless of concentration, it elicits significant dose-dependent inhibition of the viability of HT29, HCT116, and DLD1 cells with a 48 h IC ₅₀ of 6.8, 5.0, and 3.2 μM, respectively. The expression of CK1α in CRC primary cultures and PDX samples, significantly correlated with Ki-67 expression, and both were attenuated by Epiblastin A. We also observed that the effect of 5 mg/kg Epiblastin A on tumor volume, and body weight in the CRC PDX mice models, was similar to that of 5 mg/kg Cetuximab over the time-course of our in vivo study. In DLD1-derived tumor xenograft mice, Epiblastin A with very mild effect on mice body weight, suppressed tumor volume and tumor weight in a CK1α-dependent manner (P=0.024).

Conclusions

Our results demonstrate the efficacy of Epiblastin A in CRC and its potential as a putative small-molecule inhibitor of CK1α and Ki-67 signaling, which are relevant in the CRC initiation, progression and prognosis.