Rebound increase in microRNA levels at the end of 5-FU-based therapy in colorectal cancer patients

Abstract

Treatment with 5-fluorouracil (5-FU) based therapy is still used for colorectal cancer (CRC). Epigenetics has become a focus of study in cancer because of its reversibility besides its known regulatory functions. In this study, we will monitor the change in microRNAs (miRNAs) levels with 5-FU-based therapy at baseline and after 3 and 6 months of treatment to be correlated with their prognostic potential. The expression levels of 5 miRNAs, namely miRNA223-3p, miRNA20a-5p, miRNA17-5p, miRNA19a-3p, and miRNA7-5p, were measured in the peripheral blood of 77 CRC patients, along with the expression of 3 proteins PTEN, ERK, and EGFR. At baseline, CRC patients had significantly higher levels of circulating miRNAs than healthy controls. This level was reduced after 3 months of 5-FU-based therapy, then increased after 6 months significantly in responder patients compared to non-responders. MiRNA19a-3p showed that significant pattern of change in the subgroups of patients with high ERK, EGFR, and PTEN protein levels, and its 6 months level after 5-FU-based therapy showed significance for the hazard of increased risk of disease recurrence and progression.

Figure 1

Expression levels of 2^−ΔCT of miRNAs in the serum of baseline 77 CRC patients compared to 20 healthy subjects. (A) miRNA223-3p, (B) miRNA20a-5p, (C) miRNA17-5p, (D) miRNA19a-3p, (E) miRNA7-5p, (F) ROC curve showing AUC of miRNAs at baseline patients compared to the line of reference. miRNAs levels are expressed as 2^−ΔCT relative to RNU6 amplification level.

Figure 2

Change in miRNAs levels with 5-FU-based therapy in CRC patients at baseline, and after 3 and 6 months of treatment. (A) in responders (n = 23), (B) in non-responders (n = 20). I. miRNA223-3p, II. miRNA20a.5p, III. miRNA17-5p, IV. miRNA19a-3P, V. miRNA7-5p. The levels of miRNAs are expressed as log2^−ΔΔCT, and P-values are displayed in the figures.

Figure 3

Change in miRNA19a-3p level in response to three and six months of 5-FU therapy in patients with low and high basal levels of proteins. (A) EGFR, (B) ERK (C) PTEN. The levels of miRNAs are expressed as log2^−ΔΔCT, and the P-values are displayed in the figures.

Figure 4

Level of miRNAs in non-metastatic and metastatic patients after three and six months of 5-FU therapy. (A) miRNA223-3p, (B) miRNA20a-5p, (C) miRNA19a-3p, (D) miRNA17-5p, (E) miRNA7-5p. The levels of miRNAs are expressed as log2^−ΔΔCT and P-values are displayed on the figures.

Figure 5

The hazard of disease recurrence and progression after 3 years of follow-up for 77, 60 and 41 CRC patients at baseline, after 3 months of 5-FU therapy, after 6 months of 5-FU therapy, respectively. Data presented as median logHR (95% CI) of disease recurrence and progression after 3 years of follow-up. Abbreviations: HR: hazard ratio, CI: confidence interval.