CL Shapiro

Patients with recurrent breast cancer can be divided into three categories: those with locoregional recurrence, those with distant nonvisceral recurrence, and those with visceral recurrence. Survival from time of first relapse is clearly dependent on these categories. Selection of therapeutic modality should be based on considerations of site recurrence, symptomatology, anticipated response to therapy, and expected toxicities related to the therapy. Chemotherapy is appropriate for patients who are either unlikely to respond to hormone therapy, quite symptomatic, clearly hormone refractory, or have rapidly progressive visceral disease. Previously untreated patients are likely to respond to chemotherapy, with no clear-cut marker or clinical category associated with increased or decreased likelihood of benefit. Studies are ongoing to identify markers for response to chemotherapy, with recent investigations focusing on HER-2/neu expression. Standard combination chemotherapeutic regimens, consisting of either cyclophosphamide/methotrexate/5-fluorouracil (CMF) or cyclophosphamide/doxorubicin/5-fluorouracil (CAF), are associated with response rates in untreated patients of 35% to 80% and in previously treated patients of 10% to 40%. Although CAF probably has a slightly higher response rate than CMF, the toxicity of CAF is substantially higher. Newer agents are effective in both previously untreated and treated patients with breast cancer. These include paclitaxel, docetaxel, vinorelbine (Navelbine; Burroughs Wellcome Co, Research Triangle Park, NC; Pierre Fabre Médicament, Paris, France), and amonafide. Furthermore, modulation of previously existing agents, designed to overcome resistance, has been tested. Only leucovorin/5-fluorouracil has apparent clinical benefit. A number of novel approaches are being designed or are currently being used in clinical trials. These include differentiating agents, anti-angiogenesis factors, antitumor antigen-based therapy, growth factor receptor/ligand therapy, and gene therapy.

Abstract Abstract #6131 Background: Treatment options for postmenopausal women (PW) with hormone responsive breast cancer (HRBC) involve blocking estrogen receptor (ER) with anti-estrogens or inhibiting estrogen synthesis with aromatase inhibitors (AI). A mouse xenograft model showed that combination of the pure estrogen antagonist fulvestrant (FUL) and an AI was more effective in suppressing tumor growth than either treatment alone. We hypothesized this combination would have an enhanced anti-tumor activity in PW with HRBC. To test this hypothesis a phase II trial of FUL and the steroidal AI exemestane (EXE) was designed.
 Materials & Methods: Eligible PW may have received prior adjuvant chemotherapy (CT), treatment with tamoxifen, anastrazole or letrozole in the adjuvant or advanced disease setting. EXE at a dose of 25 mg daily was started on Day 1. On day 8, FUL was added and administered as an IM injection of 250 mg every 28 days. Response was assessed every 2 cycles by physical examination and imaging studies. The steady-state pharmacokinetics of EXE when administered alone (Day 7) and in combination with FUL (Day 120) was assessed in a subset of PW. Plasma samples were obtained before and 1, 2, 4, 6, 8, and 24 hours after dosing on each occasion. Plasma concentrations of EXE and 17-hydroxyEXE were analyzed using a validated liquid chromatographic method with mass spectrometric detection. Serum concentrations of insulin growth factor I (IGF-I) were determined prior to dosing, on Day 7 and Day 120. The primary clinical endpoint is the proportion of patients free of progressive disease at 6 months. A Fleming single-stage design (with P0 =0.5, P1= 0.7, α=0.10 and β=0.10) requires a sample size of 40 women. If 24 of the 40 PW are progression-free at 6-months, this treatment will be worthy of future study.
 Results: Nineteen women were enrolled. The median age was 64 years (range 48-83 years); 42% had prior adjuvant CT, 53% had prior hormonal therapy; and 47% had visceral metastases. Nine (47%) had a PFS ≥ 6 months (range 6 to 20 months) with 8 still on treatment; 8 (42%) progressed prior to 6 mo; and 2 (11%) have been on treatment for < 2 mo. Only 1 (5%) developed grade 3 arthralgias that required treatment discontinuation; 2 (11%) had grade 3 fatigue; and 2 (11%) had grade 3 bone pain. Steady-state EXE plasma concentration-time profiles were determined in 9 women on both Day 7 (EXE alone) and Day 120 (EXE+FUL). Mean EXE Cmax, AUC, and half-life values determined on Day 120 were within 12% of values determined on Day 7. The median serum concentrations of IGF-I prior to therapy were 112 ng/ml (range 85-172); on day 7 were 134 ng/ml (range 86-174); and on day 120 were 129 ng/ml (range 73-216) Discussion: Co-administration of FUL does not result in clinically relevant changes in EXE plasma concentrations. This combined therapy is well tolerated and active. Accrual is ongoing. Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 6131.

Abstract Abstract #3023 Background: Breast cancer is the most common malignancy in women, accounting for 31% of all female cancers. Over two-thirds of breast cancers exhibit high concentrations of estrogen receptor, which contribute to tumor growth and progression. Blocking the steroid hormone pathway with tamoxifen and/or oophorectomy has been shown to be effective in this patient population. However, approximately 30% of the breast cancer is resistant to tamoxifen. Recent studies have highlighted the key regulatory roles of microRNAs (miR) in all fundamental cellular processes in animals and plants including primary human cancers. We hypothesized that alteration in the expression of specific miRs in breast cancer could contribute to tamoxifen resistance.
 Methods: To test this hypothesis, we performed microRNA microarray analysis using MCF-7 cell lines that are either sensitive (parental) or resistant to tamoxifen (4-hydroxy tamoxifen resistant-OHTR). Using Real-time RT-PCR we validated altered expression of the miRs in both the cell culture model and the primary human breast cancer tissues. Cells overexpressing miR-221/222 or p27(kip1) were created by transfection of mammalian expression vectors using Lipofectamine2000 followed by G418 selection. Cell viability upon tamoxifen treatment was measured by MTT assay, extent of apoptosis was monitored by Western blot analysis of PAPR and Caspase cleavage and cell cycle profile was studied using Flow cytometry.
 Results: Eight miRs were found to be significantly upregulated while seven miRs were significantly down-regulated in the OHTR cells compared to parental MCF-7 cells. The increased expression of three upregulated (miR-221, miR-222 and miR-181) and three downregulated (miR-21, miR-342 and miR-489) miRs was later validated in the cell lines by real-time RT-PCR. In addition, the level of miR-221 and miR-222 was significantly elevated in Her2/neu(+) primary human breast cancer tissues, compared to Her2/neu(-) tissue samples. The Her2/neu expressing tumors are known to be relatively resistant to endocrine therapy. Ectopic expression of miR-221/miR-222 in parental MCF-7 cells rendered the cells more tolerant to tamoxifen than the control cells. The cell cycle inhibitor p27/Kip1, a known target of miR-221/miR-222 was significantly reduced in both OHTR cells and miR-221/222 overexpressing MCF-7 cells, which was consistent with the upregulation of the miRs. Ectopic expression of p27/Kip1 in the resistant OHTR cells enhanced cell death when exposed to tamoxifen.
 Conclusion: This study has revealed a specific miR signature for the tamoxifen-resistant breast cancer and an important cell cycle inhibitor target of the altered miR, which could be used as a prognostic marker for the drug-resistant breast cancer. Further studies of other miRs differentially expressed in tamoxifen resistant cell lines, will help us not only in identifying such patients but also may serve as a therapeutic target in the future. Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3023.

BACKGROUND: Alkylating agents administered as single agents or in combination with antimetabolites or anthracyclines delay the appearance of metastases and prolong the survival of breast cancer patients after surgery. PURPOSE: This phase III clinical trial was designed to evaluate the therapeutic efficacy and toxicity of the alkylating agent cyclophosphamide in combination with the antimetabolites methotrexate and fluorouracil adjuvant to breast cancer surgery. METHODS: This study consisted of 255 breast cancer patients (a) with one to three histologically positive axillary lymph nodes and either no detectable primary tumor or operable primary tumors 5 cm or less (T0-T2) (95% of the patients) or (b) with tumors larger than 5 cm (T3) and with negative axillary nodes. Patients were randomly allocated to receive either methotrexate (60 mg/m2) and fluorouracil (600 mg/m2) (MF) intravenously on days 1 and 8 every 28 days for eight cycles or cyclophosphamide (100 mg/m2) orally on days 1-14 plus MF (CMF) every 28 days for the same duration. Median follow-up was 7.8 years, and maximum follow-up was 13 years. RESULTS: There were no statistically significant differences in time to treatment failure or overall survival for patients treated with MF or CMF. At 8 years after completion of treatment, time to treatment failure was 55% (95% confidence interval [CI] = 50%-60%) and 59% (95% CI = 54%-64%) and overall survival was 69% (95% CI = 65%-73%) and 67% (95% CI = 62%-72%) for MF- and CMF-treated patients, respectively. The hazard ratios (MF to CMF) for time to treatment failure and for survival, estimated with a proportional hazards model, were 1.02 (95% CI = 0.69-1.50) and 0.87 (95% CI = 0.56-1.34), respectively. Myelosuppression was significantly greater (P < .0001) in CMF-treated patients during cycles 1-6. Median white blood cell count nadirs were between 4.4 x 10(3)/microL and 3.5 x 10(3)/microL in patients receiving MF and between 3.0 x 10(3)/microL and 2.4 x 10(3)/microL in those receiving CMF. Dose reductions were more frequent in CMF-treated patients, which led to statistically significant differences (P < .0001) in amounts of methotrexate and fluorouracil administered. Primary cancers at other sites occurred in 14 patients (5.5%)--six treated with MF and eight treated with CMF. CONCLUSIONS: Our findings suggest that the addition of cyclophosphamide to adjuvant chemotherapy with MF offers no therapeutic advantage but results in increased myelosuppression. IMPLICATIONS: Future trials will define the possible advantages of antimetabolites in adjuvant therapy. Further information will also become available when results of the ongoing National Surgical Adjuvant Breast and Bowel Project trial comparing adjuvant MF to CMF in node-negative breast cancer patients are presented.

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