Prognostic significance of minichromosome maintenance proteins in breast cancer

Abstract

A role for the minichromosome maintenance (MCM) proteins in cancer initiation and progression is slowly emerging. Functioning as a complex to ensure a single chromosomal replication per cell cycle, the six family members have been implicated in several neoplastic disease states, including breast cancer. Our study aim to investigate the prognostic significance of these proteins in breast cancer. We studied the expression of MCMs in various datasets and the associations of the expression with clinicopathological parameters. When considered alone, high level MCM4 overexpression was only weakly associated with shorter survival in the combined breast cancer patient cohort (n = 1441, Hazard Ratio = 1.31; 95% Confidence Interval = 1.11-1.55; p = 0.001). On the other hand, when we studied all six components of the MCM complex, we found that overexpression of all MCMs was strongly associated with shorter survival in the same cohort (n = 1441, Hazard Ratio = 1.75; 95% Confidence Interval = 1.31-2.34; p < 0.001), suggesting these MCM proteins may cooperate to promote breast cancer progression. Indeed, their expressions were significantly correlated with each other in these cohorts. In addition, we found that increasing number of overexpressed MCMs was associated with negative ER status as well as treatment response. Together, our findings are reproducible in seven independent breast cancer cohorts, with 1441 patients, and suggest that MCM profiling could potentially be used to predict response to treatment and prognosis in breast cancer patients.

Keywords: Minichromosome maintenance complex; breast cancer; prognosis; survival.

Figure 1

Differential expression of MCM4 in paired breast cancer and non-tumor breast epithelium specimens. A. mRNA expression of MCM4 determined by Real-time PCR in the 35 pairs of tumor and non-tumor specimens. B. Protein expression of MCM4 determined by immunohistochemical staining of the 45 pairs of tumor and non-tumor specimens.

Figure 2

The association between MCM4 expression and ER status of breast tumors. (A) Protein expression of MCM4 determined by immunohistochemical staining in 90 breast cancer specimens with different ER status. (B-D) Relative expression of MCM4 mRNA determined by microarray analysis in breast cancer specimens with different ER status in (B) GSE2034, (C) GSE3494 and (D) GSE7390 breast cancer datasets. (E) Percentage of cases with high or low MCM4 mRNA expression in breast cancer specimens with different ER status in the combined breast cancer dataset.

Figure 3

The association between MCM4 expression and histological grading of breast cancers. (A) Protein expression of MCM4 determined by immunohistochemical staining in 91 breast cancer specimens with different histological grading. (B-E) Relative expression of MCM4 mRNA determined by microarray analysis in breast cancer specimens with different histological grading in (B) GSE1456, (C) GSE3494, (D) GSE7390 and (E) GSE11121 breast cancer datasets.

Figure 4

The association between MCM4 expression and survival of breast cancer patients. MCM4 mRNA expression was stratified into high or low expression using the median mRNA expression level as the cut-off point. Kaplan-Meier survival curve for MCM expression and the p-value for log-rank test showed alongside for (A) GSE1456, (B) GSE2034, (C) GSE3143, (D) GSE3494, (E) GSE7390, (F) GSE11121, (G) GSE12276 and (H) for all seven breast cancer datasets combined.

Figure 5

The association between MCMs expression and survival of breast cancer patients. MCM mRNA expression was stratified into high or low expression using the median mRNA expression level as the cut-off point. Kaplan-Meier survival curve for (A) MCM2, (B) MCM3, (C) MCM5, (D) MCM6 and (E) MCM7 expression and the p-value for log-rank test showed alongside for the combined breast cancer dataset.

Figure 6

The association between a combinatorial score for MCM2-7 expression and survival of breast cancer patients. (A-G) MCM mRNA expression was stratified into high or low expression using the median mRNA expression level as the cut-off point. Specimens were than stratified into two groups by the number of MCMs expressed at a high level (≤ 3 MCMs or ≥ 4 MCMs expressed at a high level). Kaplan-Meier survival curve for a combinatorial score for MCM2-7 expression and the p-value for log-rank test showed alongside for (A) GSE1456, (B) GSE2034, (C) GSE3141, (D) GSE3494, (E) GSE7390, (F) GSE11121 and (G) GSE12276. (H) Kaplan-Meier survival curve for specimens stratified into seven groups based on the number of MCMs expressed at a high level. (I) Kaplan-Meier survival curve for specimens stratified into two groups based on the number of MCMs expressed at a high level (≤ 3 MCMs or ≥ 4 MCMs expressed at a high level).

Figure 7

The association between MCMs expression and the response to treatment. Breast cancer patients were stratified according to their response to therapeutic intervention (namely residual disease or pCR for GSE22093 (A); no response, partial response, near complete response or complete response for GSE22358 (B); no pCR or pCR for GSE42822 (C), and this response was correlated with the number of MCM genes overexpressed in individuals’ tumors.

Figure 8

The association of ER status and MCMs mRNA expression, survival and response to letrozole treatment in breast cancer patients. Kaplan-Meier survival curve for MCM4 in specimens with (A) ER-negative and (B) ER positive status. (C) Histogram showing the percentage of cases expressing different numbers of MCMs at a high level. (D) Histogram showing the percentage of cases expressing ≤ 3 MCMs or ≥ 4 MCMs at a high level. Kaplan-Meier survival curve for MCMs combinatorial score (≤ 3 MCMs or ≥ 4 MCMs expressed at a high level) in specimens with (E) ER-negative and (F) ER positive status. (G) Histogram summarizing the impact of letrozole treatment on the expression of MCM mRNAs.