Clinical significance of miR-144-ZFX axis in disseminated tumour cells in bone marrow in gastric cancer cases

BACKGROUND: We previously reported that bone marrow (BM) was a homing site for gastric cancer (GC) cells leading to haematogenous metastases. There has been little study that microRNAs regulated pathways in malignant cells or host cells in BM, and thereby regulated the progression of GC. METHODS: Both microRNA microarray and gene expression microarray analyses of total RNA from BM were conducted, comparing five early and five advanced GC patients. We focused on miR-144-ZFX axis as a candidate BM regulator of GC progression and validated the origin of the microRNA expression in diverse cell fractions (EpCAM(+)CD45(−), EpCAM(−)CD45(+), and CD14(+)) by magnetic-activated cell sorting (MACS). RESULTS: Quantitative reverse-transcriptase (RT)–PCR analysis validated diminished miR-144 expression in stage IV GC patients with respect to stage I GC patients (t-test, P=0.02), with an inverse correlation to ZFX (ANOVA, P<0.01). Luciferase reporter assays in five GC cell lines indicated their direct binding and validated by western blotting. Pre-miR144 treatment and the resultant repression of ZFX in GC cell lines moderately upregulated their susceptibility to 5-fluorouracil chemotherapy. In MACS-purified BM fractions, the level of miR-144 expression was significantly diminished in disseminated tumour cell fraction (P=0.0005). Diminished miR-144 expression in 93 cases of primary GC indicated poor prognosis. CONCLUSION: We speculate that disseminated cancer cells could survive in BM when low expression of miR-144 permits upregulation of ZFX. The regulation of the miR-144-ZFX axis in cancer cells has a key role in the indicator of the progression of GC cases.