Discovery of genes from feces correlated with colorectal cancer progression

Abstract

Colorectal cancer (CRC) is considered to develop slowly via a progressive accumulation of genetic mutations. Markers of CRC may serve to provide the basis for decision-making, and may assist in cancer prevention, detection and prognostic prediction. DNA and messenger (m)RNA molecules that are present in human feces faithfully represent CRC manifestations. In the present study, exogenous mouse cells verified the feasibility of total fecal RNA as a marker of CRC. Furthermore, five significant genes encoding solute carrier family 15, member 4 (SLC15A4), cluster of differentiation (CD)44, 3-oxoacid CoA-transferase 1 (OXCT1), placenta-specific 8 (PLAC8) and growth arrest-specific 2 (GAS2), which are differentially expressed in the feces of CRC patients, were verified in different CRC cell lines using quantitative polymerase chain reaction. The present study demonstrated that the mRNA level of SLC15A4 was increased in the majority of CRC cell lines evaluated (SW1116, LS123, Caco-2 and T84). An increased level of CD44 mRNA was only detected in an early-stage CRC cell line, SW1116, whereas OXCT1 was expressed at higher levels in the metastatic CRC cell line CC-M3. In addition, two genes, PLAC8 and GAS2, were highly expressed in the recurrent CRC cell line SW620. Genes identified in the feces of CRC patients differed according to their clinical characteristics, and this differential expression was also detected in the corresponding CRC cell lines. In conclusion, feces represent a good marker of CRC and can be interpreted through the appropriate CRC cell lines.

Keywords: 3-oxoacid CoA-transferase 1; cluster of differentiation 44; colorectal cancer; fecal RNA; growth arrest-specific 2; member 4; placenta-specific 8; serine/threonine kinase 17b; solute carrier family 15.

Figure 1.

Quantitation of mouse β-actin in human feces. Each NIH 3T3-containing fecal sample was stored at 4, −20 and −80°C for indicated periods. Day 3 equates to a 3-day storage, and day 7 equates to 7-day storage. Total RNA of NIH 3T3 cells in human feces was extracted and reverse transcribed into complementary DNA. The Cq value was used to indicate the expression level of the detected gene. NTC, non-template control; NIH, National Institutes of Health; Cq, quantification cycle.

Figure 2.

Relative mRNA levels of SLC15A4 in CRC cells. The expression of SLC15A4 (NM_145648) was quantified by quantitative polymerase chain reaction and normalized to the expression of endogenous glyceraldehyde 3-phosphate dehydrogenase (NM_002046). Normal colonic cell line, CCD-18Co; early-stage CRC cell lines, SW1116 (AJCC stage I) and LS123 (AJCC stage II); late-stage CRC cell lines, Caco-2 and T84 (AJCC stage IV). SLC15A4, solute carrier family 15, member 4; AJCC, American Joint Committee on Cancer; mRNA, messenger RNA; CRC, colorectal cancer.

Figure 3.

Relative mRNA levels of CD44 in early-stage CRC. The expression of CD44 (NM_001202555) was quantified by quantitative polymerase chain reaction and normalized to the expression of endogenous glyceraldehyde 3-phosphate dehydrogenase (NM_002046). Normal colonic cell line, CCD-18Co; early-stage CRC cell line, SW1116 (American Joint Committee on Cancer stage I). CD44, cluster of differentation 44; mRNA, messenger RNA; CRC, colorectal cancer.

Figure 4.

Relative mRNA levels of OXCT1 in metastatic CRC. The expression of OXCT1 (NM_000436) was quantified by quantitative polymerase chain reaction and normalized to the expression of endogenous glyceraldehyde 3-phosphate dehydrogenase (NM_002046). Non-metastatic CRC cell line, SW480; metastatic CRC cell line, CC-M3. mRNA, messenger RNA; CRC, colorectal cancer; OXCT1, 3-oxoacid CoA-transferase 1.

Figure 5.

Relative mRNA levels of genes in recurrent CRC. (A) GAS2 (NM_005256) and (B) PLAC8 (NM_016619) were quantified by quantitative polymerase chain reaction and normalized to the expression of endogenous glyceraldehyde 3-phosphate dehydrogenase (NM_002046). Non-recurrent CRC cell line, SW480; recurrent CRC cell line, SW620. mRNA, messenger RNA; CRC, colorectal cancer; PLAC8, placenta-specific 8; GAS2, growth arrest-specific 2.