Daisuke Takabatake

PTEN activity could be a predictive marker of trastuzumab efficacy in the treatment of ErbB2-overexpressing breast cancer

Trastuzumab is the only HER2/neu-directed therapy to have received Food and Drug Administration approval for the treatment of patients with metastatic breast cancer. The efficacy of trastuzumab depends on the HER2/neu status of the tumour and the patients prior treatment, but even when patients are selected on the basis of HER2/neu gene amplification, the single-agent response rate ranges from 12 to 30% and few patients respond to trastuzumab monotherapy. Here, we propose PTEN as a predictive biomarker for trastuzumab efficacy. Human breast cancer SKBR3 and drug-resistant SKBR3/R cells were investigated. We also examined clinical samples from patients who had been treated with trastuzumab and analysed the relationship between trastuzumab efficacy and PTEN level. The PI3K/Akt signalling pathway was observed to be highly active in the drug-resistant cells, and their level of PTEN was low. Delivery of antisense PTEN duplex siRNA significantly decreased the trastuzumab chemosensitivity of parental SKBR3 cells, and marked activation of Akt signalling pathway was also recognised. Moreover, immunohistochemical investigation revealed that trastuzumab treatment was remarkably successful in cells with elevated PTEN expression. Along with the immune-system-associated cytotoxic mechanism, several mechanisms have been proposed for the effect of trastuzumab. PTEN activity might play an important and major role in its HER2/PI3K/Akt-mediated antitumour effect, and could be a useful biomarker for predicting the efficacy of trastuzumab in the treatment of breast cancer.

Proteasome inhibitors can alter the signaling pathways and attenuate the P‐glycoprotein‐mediated multidrug resistance

Numerous signaling pathways were reported to be involved in the resistance for conventional cytotoxic drugs, although one of the main reasons is the overexpression of P‐glycoprotein (P‐gp) in multidrug resistant cancer cells. The overexpression of P‐gp has been associated with the resistance to a wide range of anticancer drugs. Doxorubicin and paclitaxel are substrates of this transporter system and have an important role for the various human malignancies. In the present study, drug‐sensitive MCF7 and multidrug resistant MCF7/ADR (characterized by overexpression of P‐gp) human breast cancer cell lines were used as an experimental model. We have found that PS341 and MG132, proteasome inhibitors, reduced the degree of the multidrug resistance (MDR) in MCF7/ADR cells. This phenomenon was accompanied by a decrease in the IC50 value of doxorubicin and paclitaxel from 55.9 ± 3.46 to 0.60 ± 0.08 μM, and from 17.61 ± 1.77 to 0.59 ± 0.12 μM, respectively. The IC50 values of sensitive cells for doxorubicin and paclitaxel were about 0.42 and 0.83 μM, respectively. The effect of PS341 and MG132 on MCF7/ADR cells was associated with a significant decrease in both protein and gene levels of P‐gp expression. Moreover, with regard to the expression of possible signal transduction pathways of mitogen‐activated protein kinase (MAPK) related to the activation of mdr1, proteasome inhibitors did significantly influence the activation of these proteins. Western blot analysis revealed that 24 hr exposure of multidrug resistant MCF7/ADR cells with proteasome inhibitors did change the levels of DNA binding activity of nuclear factor‐kappaB (NF‐kappaB), pERK1/2, c‐Jun, and p‐c‐Jun. In conclusion, we could remark that proteasome inhibitors (especially PS341) attenuate the resistance of MCF7/ADR cells for P‐gp substrate drugs of doxorubicin and paclitaxel. Several proteins are supposed to be associated with the resensitization of the cells to conventional cytotoxic drugs, although decreased activity of P‐gp is at least involved in the proteasome inhibitor‐related resensitization. And influence with MAPK pathways, which have been reported to be associated with the regulation of P‐gp, might be contributed to the resensitization brought by proteasome inhibitors.

Routine clinical use of the one-step nucleic acid amplification assay for detection of sentinel lymph node metastases in breast cancer patients: results of a multicenter study in Japan.

The objective of this study was to confirm, by means of a multicenter study conducted in Japan, the reliability and usefulness of the one‐step nucleic acid amplification (OSNA) assay in routine clinical use for sentinel lymph node biopsy (SLNB) of breast cancer patients.

Tumor inhibitory effect of gefitinib (ZD1839, Iressa) and taxane combination therapy in EGFR-overexpressing breast cancer cell lines (MCF7/ADR, MDA-MB-231)

Some kinds of breast cancer cell lines, similar to several types of solid tumors, express epidermal growth factor receptor (EGFR). However, gefitinib, an EGFR tyrosine kinase inhibitor, is not effective for all these cell lines. Similarly, taxane is effective for many of the cell lines, although some, such as the multidrug‐resistant MCF7/ADR cell line, show taxane‐resistance. Here, we examined the growth inhibitory effect of combination treatment with gefitinib and taxane on the breast cancer cell lines MDA‐MB‐231 (EGFR‐positive) and MCF7/ADR (EGFR‐ and HER2‐positive). To estimate the combined effect, a Combination Index was calculated for each cell line. The combination of gefitinib and taxane showed a strong synergistic effect on MCF7/ADR cells, but an invitro additive‐antagonistic effect on MDA‐MB‐231 cells. Similarly, the combination treatment showed a significantly increased tumor inhibitory effect on MCF7/ADR xenografts, but not on MDA‐MB‐231 xenografts. Regarding the mechanism of the synergistic effect, Western blotting analysis revealed that taxane activated the EGFR‐Akt pathway in MCF7/ADR cells but not in MDA‐MB‐231. To determine the optimal sequential administration of gefitinib and taxane for MCF7/ADR cells, we used flow cytometry to analyze the cell cycle and apoptosis; finding that taxane treatment followed by gefitinib produced a higher rate of G2 arrest and apoptosis than gefitinib treatment followed by taxane. These results suggest gefitinib overcomes the drug‐resistance of these cells, thereby increasing the effects of taxane on MCF7/ADR cells. Further, activation of the EGFR‐Akt pathway by taxane is related to this synergistic effect.

Feasibility study of docetaxel with cyclophosphamide as adjuvant chemotherapy for Japanese breast cancer patients.

OBJECTIVE The 7-year follow-up of the US oncology 9735 trial demonstrated the superiority of TC [docetaxel (DTX)/cyclophosphamide (CPA)] to doxorubicin/CPA therapy. To introduce TC therapy in Japan, the verification of the safety and tolerability is essential. We performed a collaborative prospective safety study with Okayama University to introduce TC therapy. METHODS The subjects were 53 patients aged from 33 to 67 years at intermediate risk based on the St Gallen risk classification who underwent radical surgery for primary breast cancer between August 2007 and December 2008. As post-operative adjuvant chemotherapy, four cycles of TC (DTX 75 mg/m(2) + CPA 600 mg/m(2)) were administered at 3-week intervals. Adverse events were evaluated based on National Cancer Institute-Common Terminology Criteria for Adverse Events ver. 3.0. The safety and completion rate were evaluated as the primary and secondary endpoints, respectively. RESULTS Regarding hematological toxicity, Grade (G) 4 neutropenia occurred in 71.7% and G3 in 26.4%. G3-4 leukopenia developed in 32.1% and 56.6%, respectively, G4 anemia in 1.9% and G1-2 anemia in 26.4%. Regarding non-hematological toxicity, systemic malaise, skin eruption, edema, myalgia, arthralgia and nausea were noted in most patients. The completion rate was 94.3%, dose reduction was necessary in 7.5% and granulocyte colony-stimulating factor (G-CSF) support was required in 17.0%. On comparison between patients aged 65 years or older and younger than 65 years, the completion rate, dose reduction and incidence of febrile neutropenia (FN) were higher in the elderly patients. G-CSF support was more often needed in this subgroup. CONCLUSIONS TC therapy is tolerable for Japanese patients, but attention should be paid to the development of FN and neutropenia. The completion rate was lower in the elderly patients, showing that tolerability was not necessarily favorable.

Gasless Endoscopic Thyroid and Parathyroid Surgery Using a New Retractor

Abstract Endoscopic thyroid and parathyroid surgery have now become feasible procedures, but the working space provided by the gasless technique is more limited than that of the CO2 insufflation technique. Gasless endoscopic surgery was performed in 20 patients with thyroid or parathyroid tumors. A newly designed retractor was used. Gasless endoscopic surgery was performed in all patients without conversion to conventional techniques. The recurrent laryngeal nerve was visualized and preserved in all patients. No recurrent nerve palsy was noted. The new retractor created a sufficient working space, and our results demonstrated the feasibility of this technique.

Factors Associated with Health-related Quality-of-life in Breast Cancer Survivors: Influence of the Type of Surgery

OBJECTIVE To determine if health-related quality-of-life (QOL) differences existed between breast cancer (BC) survivors receiving mastectomy and those receiving breast-conserving treatment (BCT). Factors associated with QOL in long-term BC survivors were also identified. METHODS One hundred patients who had previously undergone BC surgery and were alive without recurrence for >5 years were asked to answer the patient-administered questionnaires to assess their QOL (Functional Assessment of Cancer Therapy scale-Breast: FACT-B) and psychological distress (Hospital Anxiety and Depression Scale: HADS). Of them, 93 responded to the questionnaires affirmatively. RESULTS Although none of the QOL scores were related to the surgical procedures, statistically significant relationships were found between age and the scores of FACT-General and social/family well-being (SWB), and between the educational status and scores of SWB in univariate analyses. There was no statistically significant relationship between psychological distress and each factor examined. In multivariate analyses, significant correlations were established between scores of the FACT-BC subscale (FACT-BCS) and the type of surgery and between those on the FACT SWB subscale and age at study or educational status. Namely, patients who had undergone BCT, younger patients and patients with higher educational background scored higher QOL. CONCLUSIONS Among the BC survivors, those who underwent BCT experienced significantly but slightly better QOL than those who received mastectomy in FACT-BCS assessments. Younger patients and patients with higher educational backgrounds experienced significantly better SWB.

Usefulness of liquid-based cytology in hormone receptor analysis of breast cancer specimens

Immunohistochemical (IHC) analysis of the hormone receptor (HR) in breast cancer cytology is an important issue nowadays. Several studies have shown discrepancy in the HR status between the primary tumor and metastases. Cytology can be used for patients with metastatic disease. Although cytological assessment of HR is an excellent method, it has not been routinely used because of the difficulty in consistently preparing multiple good quality slides. Liquid-based cytology (LBC) preparation is considered as the key to resolving the aforementioned problem; however, few studies have reported the HR assessment in breast cancer using LBC. Therefore, the HR status of LBC slides from 82 breast cancers was compared with that of the corresponding surgical specimens. The HR assay in both the LBC slides and surgical specimens was conducted by IHC using an autostainer. For the IHC staining, the protocol recommended by the manufacturer for paraffin-embedded sections was used for both the cytology and histology specimens. The HR results of the cytology agreed with those of the histology in 80 of the 82 cases (accuracy rate, 98%) for estrogen receptor, and in 78 of the 82 cases (accuracy rate, 95%) for progesterone receptor. The overall accuracy of the HR status on the cytology and the histology was 99% in 81 of the 82 cases. In conclusion, in HR analysis of breast cancers, LBC followed by IHC using an autostainer was useful for the standard processing of cytological specimens and showed a good correlation with the results of analysis on the histology specimens.

Endometrial Metastasis from Breast Cancer during Adjuvant Endocrine Therapy

It is well-known that tamoxifen increases the risk of endometrial cancer. Although metastasis to the uterus from breast cancer is uncommon, there have been some case reports on uterine metastasis. If an endometrial abnormality is detected, the differential diagnosis of whether the uterine tumor is metastatic or primary is very important to determine the course of treatment. We herein report a case in which we detected a uterine tumor during follow-up after treatment with tamoxifen, and demonstrate that GCDFP-15 is useful in diagnosing metastatic uterine tumors arising from breast cancer.

Phase I and pharmacokinetic study of trastuzumab emtansine in Japanese patients with HER2-positive metastatic breast cancer

OBJECTIVE Trastuzumab emtansine (T-DM1), an antibody-drug conjugate composed of the cytotoxic agent DM1 conjugated to trastuzumab via a stable thioether linker, has shown clinical activity in human epidermal growth factor receptor 2-positive metastatic breast cancer patients. This study evaluated the maximum tolerated dose, toxicity and pharmacokinetics of trastuzumab emtansine in Japanese breast cancer patients. METHODS Inoperable advanced or recurrent human epidermal growth factor receptor 2-positive breast cancer patients were administered trastuzumab emtansine intravenously at a dose of 1.8, 2.4 or 3.6 mg/kg every 3 weeks. The maximum tolerated dose was estimated using the continual reassessment method. RESULTS This study enrolled 10 patients who were administered trastuzumab emtansine for a median of seven cycles. The dose-limiting toxicity was Grade 3 elevation of aspartate aminotransferase/alanine aminotransferase at the 2.4 mg/kg dose level. The maximum tolerated dose was estimated to be 3.6 mg/kg because at the point when dose-limiting toxicity was evaluable in 10 patients, the probability of dose-limiting toxicity estimated using the continual reassessment method was closest to 25% at a dose of 3.6 mg/kg and this was unchanged by the results for patients enrolled after that. The most frequent adverse events were nausea, arthralgia, fever, fatigue and decreased appetite. Adverse events were generally tolerable. The maximum concentration and area under the concentration-time curve increased linearly with the dose. CONCLUSIONS Trastuzumab emtansine up to 3.6 mg/kg was well tolerated by Japanese breast cancer patients. Although thrombocytopenia and hepatotoxicity tended to be more severe than was seen in Western patients in previous trastuzumab emtansine trials, those adverse events recovered without special supportive treatment.