Mitsunori Sasa

Significant Effect of Polymorphisms in CYP2D6 and ABCC2 on Clinical Outcomes of Adjuvant Tamoxifen Therapy for Breast Cancer Patients

PURPOSE The clinical efficacy of tamoxifen is suspected to be influenced by the activity of drug-metabolizing enzymes and transporters involved in the formation, metabolism, and elimination of its active forms. We investigated relationships of polymorphisms in transporter genes and CYP2D6 to clinical outcome of patients receiving tamoxifen. PATIENTS AND METHODS We studied 282 patients with hormone receptor-positive, invasive breast cancer receiving tamoxifen monotherapy, including 67 patients who have been previously reported. We investigated the effects of allelic variants of CYP2D6 and haplotype-tagging single nucleotide polymorphisms (tag-SNPs) of ABCB1, ABCC2, and ABCG2 on recurrence-free survival using the Kaplan-Meier method and Cox regression analysis. Plasma concentrations of tamoxifen metabolites were measured in 98 patients receiving tamoxifen 20 mg/d. RESULTS CYP2D6 variants were significantly associated with shorter recurrence-free survival (P = .000036; hazard ratio [HR] = 9.52; 95% CI, 2.79 to 32.45 in patients with two variant alleles v patients without variant alleles). Among 51 tag-SNPs in transporter genes, a significant association was found at rs3740065 in ABCC2 (P = .00017; HR = 10.64; 95% CI, 1.44 to 78.88 in patients with AA v GG genotypes). The number of risk alleles of CYP2D6 and ABCC2 showed cumulative effects on recurrence-free survival (P = .000000055). Patients carrying four risk alleles had 45.25-fold higher risk compared with patients with <or= one risk allele. CYP2D6 variants were associated with lower plasma levels of endoxifen and 4-hydroxytamoxifen (P = .0000043 and .00052), whereas no significant difference was found among ABCC2 genotype groups. CONCLUSION Our results suggest that polymorphisms in CYP2D6 and ABCC2 are important predictors for the prognosis of patients with breast cancer treated with tamoxifen.

Impact of CYP2D6*10 on recurrence‐free survival in breast cancer patients receiving adjuvant tamoxifen therapy

The clinical outcomes of breast cancer patients treated with tamoxifen may be influenced by the activity of cytochrome P450 2D6 (CYP2D6) enzyme because tamixifen is metabolized by CYP2D6 to its active forms of antiestrogenic metabolite, 4‐hydroxytamoxifen and endoxifen. We investigated the predictive value of the CYP2D6*10 allele, which decreased CYP2D6 activity, for clinical outcomes of patients that received adjuvant tamoxifen monotherapy after surgical operation on breast cancer. Among 67 patients examined, those homozygous for the CYP2D6*10 alleles revealed a significantly higher incidence of recurrence within 10 years after the operation (P = 0.0057; odds ratio, 16.63; 95% confidence interval, 1.75–158.12), compared with those homozygous for the wild‐type CYP2D6*1 alleles. The elevated risk of recurrence seemed to be dependent on the number of CYP2D6*10 alleles (P = 0.0031 for trend). Cox proportional hazard analysis demonstrated that the CYP2D6 genotype and tumor size were independent factors affecting recurrence‐free survival. Patients with the CYP2D6*10/*10 genotype showed a significantly shorter recurrence‐free survival period (P = 0.036; adjusted hazard ratio, 10.04; 95% confidence interval, 1.17–86.27) compared to patients with CYP2D6*1/*1 after adjustment of other prognosis factors. The present study suggests that the CYP2D6 genotype should be considered when selecting adjuvant hormonal therapy for breast cancer patients. (Cancer Sci 2008; 99: 995–999)

A dose‐response relationship between the frequency of p53 mutations and tobacco consumption in lung cancer patients

Mutations of the p53 tumor suppressor gene are frequent in lung cancers. It is suggested that p53 mutations are associated with smoking‐induced lung carcinogenesis. We examined p53 mutations in 53 lung cancers by analyzing reverse transcription‐polymerase chain reaction‐single strand conformation polymorphism (RT‐PCR‐SSCP) to ascertain the association between p53 mutations and smoking. Twenty‐five (47%) of 53 lung cancers carried p53 mutations. A discriminant analysis showed that the Brinkman index (0.156) and gender (0.140) significantly influenced p53 mutations. Furthermore, there was a dose‐response relationship between the quantity of cigarettes consumed and the frequency of p53 mutations in lung cancer patients (P < 0.001). In patients with adenocarcinoma, the frequency of p53 mutations correlated with the amount of the tobacco smoked (P < 0.05). We suggest that the p53 gene is a target of particular carcinogen in tobacco smoke.

Human papillomavirus type 33 dna in breast cancer in Chinese

BackgroundThe association between human papillomavirus (HPV) and anogenital tumors, especially cervical cancer, is well documented. However, it remains unclear whether there is also a correlation between HPV infection and human breast cancer.MethodsWe used PCR and Southern blot hybridization to analyze HPV-related DNA specimens from 32 cases of invasive ductal carcinoma operated upon in the Shanghai region of China.ResultsDNA derived from HPV33 was detected in 14 cases (43.8%). No HPV16 or HPV18 DNA was detected in any of the cases in this study. This is the first report demonstrating a correlation between HPV33 infection and breast cancer.ConclusionsOur results suggest that HPV33 infection may be involved in the pathogenesis of breast cancer in Chinese.

A genome-wide association study identifies locus at 10q22 associated with clinical outcomes of adjuvant tamoxifen therapy for breast cancer patients in Japanese

Although many association studies of polymorphisms in candidate genes with the clinical outcomes of breast cancer patients receiving adjuvant tamoxifen therapy have been reported, genetic factors determining individual response to tamoxifen are not fully understood. To identify genetic polymorphisms associated with clinical outcomes of patients with tamoxifen treatment, we conducted a genome-wide association study (GWAS). We studied 462 Japanese patients with hormone receptor-positive, invasive breast cancer receiving adjuvant tamoxifen therapy. Of them, 240 patients were analyzed by genome-wide genotyping using the Illumina Human610-Quad BeadChips, and two independent sets of 105 and 117 cases were used for replication studies. In the GWAS, we detected significant associations with recurrence-free survival at 15 single-nucleotide polymorphisms (SNPs) on nine chromosomal loci (1p31, 1q41, 5q33, 7p11, 10q22, 12q13, 13q22, 18q12 and 19p13) that satisfied a genome-wide significant threshold (log-rank P= 2.87 × 10(-9)-9.41 × 10(-8)). Among them, rs10509373 in C10orf11 gene on 10q22 was significantly associated with recurrence-free survival in the replication study (log-rank P= 2.02 × 10(-4)) and a combined analysis indicated a strong association of this SNP with recurrence-free survival in breast cancer patients treated with tamoxifen (log-rank P= 1.26 × 10(-10)). Hazard ratio per C allele of rs10509373 was 4.51 [95% confidence interval (CI), 2.72-7.51; P= 6.29 × 10(-9)]. In a combined analysis of rs10509373 genotype with previously identified genetic makers, CYP2D6 and ABCC2, the number of risk alleles of these three genes had cumulative effects on recurrence-free survival among 345 patients receiving tamoxifen monotherapy (log-rank P= 2.28 × 10(-12)). In conclusion, we identified a novel locus associated with recurrence-free survival in Japanese breast cancer patients receiving adjuvant tamoxifen therapy.

3-dimensional computed tomography lymphography-guided identification of sentinel lymph nodes in breast cancer patients using subcutaneous injection of nonionic contrast medium: a clinical trial.

Objective: Simple and reliable identification methods for sentinel lymph nodes (SLNs) which do not use radioisotope are essential for early breast cancer patients in community hospitals in Japan. The purpose of this paper is to demonstrate the feasibility and efficacy of computed tomography (CT) lymphography for SLN detection. Methods: The study included 15 cases with T1 or T2 breast cancer. After subcutaneous injection of 1 mL of iopamidol in 1 subareolar area of the affected breast, CT scanning was carried out and 3-dimensional (3D) CT images were created. SLNs predicted from images and CT values were assessed as to whether they were identical to those identified by the dye method. Results: An enhanced lymph vessel draining into SLN was demonstrated in 11 cases (73%) and an enhanced SLN in 10 cases (67%). 3D images clearly revealed the anatomic relationship between lymph vessels, SLN, and the surrounding structures. In addition, SLN could be predicted by the change of CT value in the time-course in another case. In total, SLN in 13 cases (87%) could be predicted. All SLNs suggested from CT lymphography were identified by the dye method. No significant adverse effect was noted in any case. Conclusions: The present clinical trial indicated that subcutaneous injection of nonionic contrast medium with CT scanning seems to be a promising method for the demonstration of a draining lymph vessel and SLN. The CT value time-course may also provide some important information. Further trials will be needed for the successful establishment of this CT lymphography–guided method for SLN identification.

Lessons for pharmacogenomics studies: association study between CYP2D6 genotype and tamoxifen response.

We earlier reported a significant association between the cytochrome P450 2D6 (CYP2D6) genotype and the clinical outcome in 282 Japanese breast cancer patients receiving tamoxifen monotherapy. Although many research groups have provided evidence indicating the CYP2D6 genotype as one of the strongest predictors of tamoxifen response, the results still remain controversial. We hypothesized that concomitant treatment was one of the causes of these controversial results. We then studied 167 breast cancer patients who received tamoxifen-combined therapy to evaluate the effects of concomitant treatment on the association analysis and observed no significant association between CYP2D6 genotype and recurrence-free survival (P=0.44, hazard ratio: 0.64, 95% confidential interval: 0.20-1.99 in patients with two variant alleles vs. patients without a variant allele). When we carried out two subgroup analyses for nodal status and tumor size, we observed a positive association between the CYP2D6 genotype and the clinical outcome only in patients who received tamoxifen monotherapy. This study explained a part of the discrepancies among the reported results.

Molecular features of triple negative breast cancer cells by genome-wide gene expression profiling analysis

Triple negative breast cancer (TNBC) has a poor outcome due to the lack of beneficial therapeutic targets. To clarify the molecular mechanisms involved in the carcinogenesis of TNBC and to identify target molecules for novel anticancer drugs, we analyzed the gene expression profiles of 30 TNBCs as well as 13 normal epithelial ductal cells that were purified by laser-microbeam microdissection. We identified 301 and 321 transcripts that were significantly upregulated and downregulated in TNBC, respectively. In particular, gene expression profile analyses of normal human vital organs allowed us to identify 104 cancer-specific genes, including those involved in breast carcinogenesis such as NEK2, PBK and MELK. Moreover, gene annotation enrichment analysis revealed prominent gene subsets involved in the cell cycle, especially mitosis. Therefore, we focused on cell cycle regulators, asp (abnormal spindle) homolog, microcephaly-associated (Drosophila) (ASPM) and centromere protein K (CENPK) as novel therapeutic targets for TNBC. Small-interfering RNA-mediated knockdown of their expression significantly attenuated TNBC cell viability due to G1 and G2/M cell cycle arrest. Our data will provide a better understanding of the carcinogenesis of TNBC and could contribute to the development of molecular targets as a treatment for TNBC patients.

Clinical efficacy and problems with CT lymphography in identifying the sentinel node in breast cancer

BackgroundCombining a radioisotope with a dye-guided method is the best method for identification of the sentinel lymph nodes (SNs) in breast cancer. However, some institutions are limited to use of a dye-guided method alone. Recently, computed tomographic lymphography (CTLG) employing a nonionic contrast medium has achieved SN identification.Patients and methods218 patients with primary breast cancer and no clinical evidence of lymph node metastasis were studied. SN identification was performed by CTLG and a dye-guided method. The SN identification rate was analyzed for correlations with the clinicopathological findings.ResultsThe SN identification rates were 96% with CTLG, 92% with the dye-guided method and 99% with both methods combined. The identification rates with CTLG and the combined method were significantly lower in node-positive patients compared to node-negative patients, and significantly lower with the combined method in vascular invasion-positive patients compared to negative patients. In addition, the SN identification rate with the dye-guided method was significantly lower in patients with a body mass index (BMI) of ≥ 25, whereas the BMI did not affect the identification rate with CTLG or the combined method. Multiple SNs were detected in approximately 20% of the patients.ConclusionCombined performance of CTLG and a dye-guided method enables identification of SNs prior to breast cancer surgery. That SN identification is easier compared with by the dye-guided method alone, and the identification rate is improved compared with either method alone. The combination of methods was especially useful in obese patients. For patients with multiple SNs, the combination has the further advantage of enabling accurate SN biopsy. CTLG may yield false-negative findings in node-positive patients and patients with lymph vessel obstruction.

Targeting BIG3–PHB2 interaction to overcome tamoxifen resistance in breast cancer cells

The acquisition of endocrine resistance is a common obstacle in endocrine therapy of patients with oestrogen receptor-α (ERα)-positive breast tumours. We previously demonstrated that the BIG3–PHB2 complex has a crucial role in the modulation of oestrogen/ERα signalling in breast cancer cells. Here we report a cell-permeable peptide inhibitor, called ERAP, that regulates multiple ERα-signalling pathways associated with tamoxifen resistance in breast cancer cells by inhibiting the interaction between BIG3 and PHB2. Intrinsic PHB2 released from BIG3 by ERAP directly binds to both nuclear- and membrane-associated ERα, which leads to the inhibition of multiple ERα-signalling pathways, including genomic and non-genomic ERα activation and ERα phosphorylation, and the growth of ERα-positive breast cancer cells both in vitro and in vivo. More importantly, ERAP treatment suppresses tamoxifen resistance and enhances tamoxifen responsiveness in ERα-positive breast cancer cells. These findings suggest inhibiting the interaction between BIG3 and PHB2 may be a new therapeutic strategy for the treatment of luminal-type breast cancer.