原発性乳癌におけるCDO1遺伝子メチル化と臨床病理学的因子の検討

2016(平成28)年

Prognostic Significance of Promoter DNA Hypermethylation of cysteine dioxygenase 1 (CDO1) Gene in Primary Breast Cancer.

Using pharmacological unmasking microarray, we identified promoter DNA methylation of cysteine dioxygenase 1 (CDO1) gene in human cancer. In this study, we assessed the clinicopathological significance of CDO1 methylation in primary breast cancer (BC) with no prior chemotherapy. The CDO1 DNA methylation was quantified by TaqMan methylation specific PCR (Q-MSP) in 7 BC cell lines and 172 primary BC patients with no prior chemotherapy. Promoter DNA of the CDO1 gene was hypermethylated in 6 BC cell lines except SK-BR3, and CDO1 gene expression was all silenced at mRNA level in the 7 BC cell lines. Quantification of CDO1 methylation was developed using Q-MSP, and assessed in primary BC. Among the clinicopathologic factors, CDO1 methylation level was not statistically significantly associated with any prognostic factors. The log-rank plot analysis elucidated that the higher methylation the tumors harbored, the poorer prognosis the patients exhibited. Using the median value of 58.0 as a cut-off one, disease specific survival in BC patients with CDO1 hypermethylation showed significantly poorer prognosis than those with hypomethylation (p = 0.004). Multivariate Cox proportional hazards model identified that CDO1 hypermethylation was prognostic factor as well as Ki-67 and hormone receptor status. The most intriguingly, CDO1 hypermethylation was of robust prognostic relevance in triple negative BC (p = 0.007). Promoter DNA methylation of CDO1 gene was robust prognostic indicator in primary BC patients with no prior chemotherapy. Prognostic relevance of the CDO1 promoter DNA methylation is worthy of being paid attention in triple negative BC cancer

BRCAness and Prognosis in Triple-Negative Breast Cancer Patients Treated with Neoadjuvant Chemotherapy.

BRCAness is defined as the set of traits in which BRCA1 dysfunction, arising from gene mutation, methylation or deletion, results in DNA repair deficiency. In the present study, we addressed BRCAness, therapeutic efficacy, recurrence, and survival in patients with triple negative breast cancer (TNBC) who were treated with neoadjuvant chemotherapy at Kitasato University Hospital, Japan, between April 2006 and October 2012. BRCAness was determined by preoperative core needle biopsy (CNB) specimens and surgical specimens. Assay was performed using Multiplex Ligation-dependent Probe Amplification (MLPA) with P376-B2 BRCA1ness probemix (MRC-Holland, Amsterdam, The Netherlands). The relative copy number ratio of each sample was compared to Human Genomic DNA (Promega, Madison, WI, USA) as reference samples was calculated with Coffalyser.NET default settings. The BRCAness score was calculated with the relative copy number ratio of various DNA sequences. Values of 0.5 or more were determined as the BRCA1-like Type (BRCAness) and those of less than 0.5 as the Sporadic Type to analyze pathological complete response (pCR) rate, recurrence, and survival. pCR (ypT0/Tis/N0) was observed in 15 patients (pCR rate: 37.5%). These patients had no recurrence. Twelve patients recurred, 8 died from breast cancer. The BRCA1-like Type were 22 and Sporadic Type were 18 in CNB specimens. No major differences were observed between the BRCA1-like Type and Sporadic Type with pCR rate, recurrence rate and survival. Twenty four surgical specimens of non-pCR patients were available and 9 were BRCA1-like Type, who had more recurrences (7/9 vs. 5/15), and their relapse-free survival was also lower (p<0.05) than that of Sporadic Type. Seven BRCA1-like Type patients remained BRCA1-like Type in surgical specimens, were worse in recurrence (p<0.01) and survival (p<0.05) compared with 6 patients whose BRCA status in surgical specimens turned to Sporadic Type. New clinical trials assessing the true recurrence (TR) rate of BRCA-type patients are expected since neither platinum-containing drugs nor poly (ADP-ribose) polymerase (PARP) inhibitors are effective against tumors with nonfunctional BRCA genes

Changes in BRCAness Types and Recurrence.

<p>Recurrent patients were shown as pink and green means no recurrence. On 40 of CNB specimens, (a) 22 patients were found to be of BRCA1-like type and (b) 18 were sporadic type. (a) BRCA1-like type; Eight patients (36.4%) achieved pCR. None of pCR patients had a recurrence. Patients whose tumor status was found to remain as BRCA1-like type upon analysis of surgical specimens experienced more recurrences than those who changed to sporadic type. (83.3% vs. 16.7% retrospectively) (p<0.01) * One patient who did not achieve pCR, residual cancer was present only in the lymph node, thus rendering the measurement of the tumor impossible. (b) Sporadic type; Seven patients (38%) achieved pCR. None of pCR patients had a recurrence. Most non-pCR patients remained sporadic type and such patients recurred at 44% (4/9).</p

Quantitative assessment of <i>CDO1</i> methylation in primary BC tissues.

<p><b>A,</b> TaqMeth value of 172 BC tissues.Median is median 58.0 (0–351.1). B, p value and relative risk were plotted according to the Log rank test. Note that the p value is constantly below 0.05, suggesting that the higher methylation value of <i>CDO1</i> gene is, poorer prognosis the patients exhibited in primary BC.</p

Characteristics of patients.

<p>Characteristics of patients.</p

BRCAness and Characters.

<p>BRCAness and Characters.</p

<i>CDO1</i> methylation and expression in BC cell line.

<p><b>A,</b><i>CDO1</i> mRNA expression in BC cell lines was assessed by semi-quantitative reverse transcribed PCR (RT-PCR). B, Representative direct bisulfite sequence results in CRL cells (methylation) and SK-BR3 cells (unmethylation). C, <i>CDO1</i> mRNA expression in BC cell lines was assessed by Q-MSP. (D) <i>CDO1</i> mRNA expression in BC tissues was assessed by RT-PCR.</p

Association between BRCAness of Surgical Specimens and Recurrence/Survival.

<p>Among 40 patients, 15 patients achieve to pCR and 1 patient had cancer on only lymph node. Twenty four breast tumors were assessed by MLPA. The recurrent rate of BRCA1-like type was worse than that of sporadic type significantly. But, survival of two groups were similar.</p