Abnormal expression of TRIB3 in colorectal cancer: a novel marker for prognosis

Abstract

Background: TRIB3 is a human homologue of Drosophila tribbles. Previous studies have shown that TRIB3 controls the cell growth through ubiquitination-dependent degradation of other proteins, whereas its significance in the prognosis of colorectal cancer (CRC) is not yet fully understood.

Materials: This study comprised 202 patients who underwent surgery for CRC, as well as 22 cell lines derived from human gastrointestinal cancer. The correlation of gene expression with clinical parameters in patients was assessed. The biological significance was evaluated by knockdown experiments in seven colorectal cancer cell lines.

Results: A total of 20 cancer cell lines (90.9%) expressed the TRIB3 gene. The assessment in surgical specimens indicated that the gene expression was significantly higher in the cancerous region than in the marginal non-cancerous region. Patients with high TRIB3 expression were statistically susceptible to a recurrence of the disease, and showed poorer overall survival than those with low expression. The assessment of TRIB3 knockdown in five cell lines showed that small interfering RNA (siRNA) inhibition resulted in a statistically significant reduction in cell growth.

Conclusion: These data strongly suggest the usefulness of TRIB3 as a marker for predicting the prognosis of CRC patients, showing a basis for the development of effective treatments for CRC.

Figure 1

TRIB3 mRNA expression in clinical tissue specimens. Quantitative real-time RT–PCR on 202 paired clinical samples showed that 181 of 202 (89.6%) samples had higher levels of TRIB3 mRNA in tumours than in paired normal regions. The mean expression value of TRIB3 mRNA in tumour regions, 154.62±1021.63 (mean±s.d.; normalised by GAPDH gene expression), was significantly higher than the value, 6.98±4.91, for the corresponding normal regions (P<0.001; Student's t-test). GAPDH=glyceraldehydes-3-phosphate dehydrogenase; RT–PCR= reverse transcriptase PCR; TRIB3=tribbles homologue 3.

Figure 2

Immunohistochemical staining for Trib3 in tumour and normal specimens. A representative positive stain for Trib3 in tissues from CRC patients. Positive staining is observed in the nucleus and cytoplasm of cancer cells, but not in stromal cells. Trib3 expression was associated with mRNA expression. CRC=colorectal cancer; T=tumour cells; N=normal glandular cells. Bar=200 μm (original magnification, × 20).

Figure 3

Overall survival rates of patients with CRC on the basis of TRIB3 mRNA expression status. The overall survival rate was significantly lower in the TRIB3 high-expression group than that in the low-expression group (P<0.001). CRC=colorectal cancer; TRIB3=tribbles homologue 3.

Figure 4

Metachronous metastasis-free over 5 years’ survival rates of patients with CRC in stages I II, and III, on the basis of TRIB3 mRNA expression status. The metachronous metastasis-free over 5 years’ survival rate was significantly lower in patients with the TRIB3 high-expression group compared with the low-expression group (P=0.007). CRC=colorectal cancer; TRIB3=tribbles homologue 3.

Figure 5

Proliferation assay with siRNA inhibition in five CRC cell lines. Proliferation assay was performed in five CRC cell lines (A, DLD-1; B, LoVo; C, HCT-116; D, KM12SM; E, SW480). There were significant differences between WT or NC, and TRIB3 siRNA. Values are presented means±s.d. of three independent experiments. CRC=colorectal cancer; NC=negative control; TRIB3=tribbles homologue 3; WT=wild type.