Noriaki Yamamoto

GSTP1 expression predicts poor pathological complete response to neoadjuvant chemotherapy in ER-negative breast cancer

The purpose of the present study was to investigate the association of glutathione S‐transferase P1 (GSTP1) expression with resistance to neoadjuvant paclitaxel followed by 5‐fluorouracil/epirubicin/cyclophosphamide (P‐FEC) in human breast cancers. The relationship of GSTP1 expression and GSTP1 promoter hypermethylation with intrinsic subtypes was also investigated. In this study, primary breast cancer patients (n = 123, stage II–III) treated with neoadjuvant P‐FEC were analyzed. Tumor samples were obtained by vacuum‐assisted core biopsy before P‐FEC. GSTP1 expression was determined using immunohistochemistry, GSTP1 promoter methylation index (MI) using bisulfite methylation assay and intrinsic subtypes using DNA microarray. The pathological complete response (pCR) rate was significantly higher in GSTP1‐negative tumors (80.0%) than GSTP1‐positive tumors (30.6%) (P = 0.009) among estrogen receptor (ER)‐negative tumors but not among ER‐positive tumors (P = 0.267). Multivariate analysis showed that GSTP1 was the only predictive factor for pCR (P = 0.013) among ER‐negative tumors. Luminal A, luminal B and HER2‐enriched tumors showed a significantly lower GSTP1 positivity than basal‐like tumors (P = 0.002, P < 0.001 and P = 0.009, respectively), while luminal A, luminal B and HER2‐enriched tumors showed a higher GSTP1 MI than basal‐like tumors (P = 0.076, P < 0.001 and P < 0.001, respectively). In conclusion, these results suggest the possibility that GSTP1 expression can predict pathological response to P‐FEC in ER‐negative tumors but not in ER‐positive tumors. Additionally, GSTP1 promoter hypermethylation might be implicated more importantly in the pathogenesis of luminal A, luminal B and HER2‐enriched tumors than basal‐like tumors. (Cancer Sci 2012; 103: 913–920)

Methylated DNA and Total DNA in Serum Detected by One-Step Methylation-Specific PCR Is Predictive of Poor Prognosis for Breast Cancer Patients

Purpose: We recently developed the one-step methylation-specific PCR (OS-MSP) assay which can detect methylated DNA (met-DNA) in serum with high sensitivity. To examine its prognostic value, we applied this new assay to the detection of met-DNA in serum of breast cancer patients. Methods: Serum samples taken before surgery from 336 primary invasive breast cancer patients were subjected to the OS-MSP assay for the promoter regions of GSTP1, RASSF1A, and RARβ2. The assay outcome was considered positive when methylation was detected in at least one of these three genes. Total DNA content in serum was also determined. Results: Of the 336 stage I/II patients, 33 (10%) were positive for met-DNA in serum and showed a significantly worse overall survival (OS) rate at 100 months (78 vs. 95%; p = 0.002) than those with negative findings (n = 303). Patients with high total DNA in serum (n = 112) also showed a significantly worse OS rate at 100 months (86 vs. 97%; p = 0.001) than those with low total DNA in serum (n = 224). Moreover, patients both positive for met-DNA and with high total DNA in serum (n = 18) showed a much worse OS rate at 100 months (65 vs. 94%; p < 0.001) than the others (n = 318). Conclusions: Met-DNA in serum detected with the OS-MSP assay constitutes a significant and independent prognostic factor, and its combination with total DNA in serum seems to be even more effective for prediction of prognosis for breast cancer patients.

Methylated DNA and high total DNA levels in the serum of patients with breast cancer following neoadjuvant chemotherapy are predictive of a poor prognosis

In a previous study, we established a one-step methylation-specific polymerase chain reaction (OS-MSP) assay for the detection of methylated DNA (met-DNA) and total DNA levels in serum. For the present study, this OS-MSP assay was used for patients with breast cancer treated with neoadjuvant chemotherapy (NAC) in order to investigate the prognostic significance of met-DNA and total DNA levels. Following treatment with NAC and prior to surgery, serum samples obtained from 120 patients with stage II/III breast cancer were subjected to the OS-MSP assay for analysis of the glutathione S-transferase pi 1, Ras association (RalGDS/AF-6) domain family member 1 and retinoic acid receptor β2 genes. The detection of methylation in a minimum of one of these genes indicated a positive outcome of the assay. The total DNA content of the serum was also determined. Of the 120 stage II/III patients, seven (6%) were positive for met-DNA in serum and showed a significantly worse overall survival (OS) time compared with patients negative for met-DNA (n=113) (5-year OS, 43 vs. 85%; P=0.002). The patients with high total DNA levels in serum (n=40) also showed a significantly worse OS compared with those with low total DNA levels (n=80) (65 vs. 91%; P<0.001). The presence of met-DNA and high total DNA levels in the serum were found to be significant prognostic factors that are independent of a pathological complete response by multivariate analysis. Following NAC, met-DNA and high total DNA levels in the serum detected with the OS-MSP assay constitute novel prognostic factors for patients with breast cancer; this may be clinically useful for the prognosis prediction for patients who do not achieve a pathological complete response following NAC.