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Abstract Introduction The detection of circulating tumour cells (CTCs) in the peripheral blood and disseminated tumour cells (DTCs) in the bone marrow are promising prognostic tools for risk stratification in early breast cancer. There is, however, a need for further validation of these techniques in larger patient cohorts with adequate follow-up periods. Methods We assayed CTCs and DTCs at primary surgery in 733 stage I or II breast cancer patients with a median follow-up time of 7.6 years. CTCs were detected in samples of peripheral blood mononuclear cells previously stored in liquid-nitrogen using a previously-developed multi-marker quantitative PCR (QPCR)-based assay. DTCs were detected in bone marrow samples by immunocytochemical analysis using anti-cytokeratin antibodies. Results CTCs were detected in 7.9% of patients, while DTCs were found in 11.7%. Both CTC and DTC positivity predicted poor metastasis-free survival (MFS) and breast cancer-specific survival (BCSS); MFS hazard ratio (HR) = 2.4 (P < 0.001)/1.9 (P = 0.006), and BCSS HR = 2.5 (P < 0.001)/2.3 (P = 0.01), for CTC/DTC status, respectively). Multivariate analyses demonstrated that CTC status was an independent prognostic variable for both MFS and BCSS. CTC status also identified a subset of patients with significantly poorer outcome among low-risk node negative patients that did not receive adjuvant systemic therapy (MFS HR 2.3 (P = 0.039), BCSS HR 2.9 (P = 0.017)). Using both tests provided increased prognostic information and indicated different relevance within biologically dissimilar breast cancer subtypes. Conclusions These results support the use of CTC analysis in early breast cancer to generate clinically useful prognostic information.

Abstract Background Presence of disseminated tumor cells (DTCs) in bone marrow (BM) after completion of systemic adjuvant treatment predicts reduced survival in breast cancer. The present study explores the use of DTCs to identify adjuvant insufficiently treated patients to be offered secondary adjuvant treatment intervention, and as a surrogate marker for therapy response. Methods A total of 1121 patients with pN1-3 or pT1c/T2G2-3pN0-status were enrolled. All had completed primary surgery and received 6 cycles of anthracycline-containing chemotherapy. BM-aspiration was performed 8-12 weeks after chemotherapy (BM1), followed by a second BM-aspiration 6 months later (BM2). DTC-status was determined by morphological evaluation of immunocytochemically detected cytokeratin-positive cells. If DTCs were present at BM2, docetaxel (100 mg/m2, 3qw, 6 courses) was administered, followed by DTC-analysis 1 month (BM3) and 13 months (BM4) after the last docetaxel infusion. Results Clinical follow-up (FU) is still ongoing. Here, the descriptive data from the study are presented. Of 1085 patients with a reported DTC result at both BM1 and BM2, 94 patients (8.7%) were BM1 positive and 83 (7.6%) were BM2 positive. The concordance between BM1 and BM2 was 86.5%. Both at BM1 and BM2 DTC-status was significantly associated with lobular carcinomas (p = 0.02 and p = 0.03, respectively; chi-square). In addition, DTC-status at BM2 was also associated with pN-status (p = 0.009) and pT-status (p = 0.03). At BM1 28.8% and 12.8% of the DTC-positive patients had ≥2 DTCs and ≥3 DTCs, respectively. At BM2, the corresponding frequencies were 47.0% and 25.3%. Of 72 docetaxel-treated patients analyzed at BM3 and/or BM4, only 15 (20.8%) had persistent DTCs. Of 17 patients with ≥3 DTCs before docetaxel treatment, 12 patients turned negative after treatment (70.6%). The change to DTC-negativity was associated with the presence of ductal carcinoma (p = 0.009). Conclusions After docetaxel treatment, the majority of patients experienced disappearance of DTCs. As this is not a randomized trial, the results can be due to effects of adjuvant (docetaxel/endocrine/trastuzumab) treatment and/or limitations of the methodology. The clinical significance of these results awaits mature FU data, but indicates a possibility for clinical use of DTC-status as a residual disease-monitoring tool and as a surrogate marker of treatment response. Trial registration Clin Trials Gov NCT00248703

Low steady‐state levels of active tamoxifen metabolites have been associated with inferior treatment outcomes. In this retrospective analysis of 406 estrogen receptor‐positive breast cancer (BC) patients receiving adjuvant tamoxifen as initial treatment, we have associated our previously reported thresholds for the two active metabolites, Z‐endoxifen and Z‐4‐hydroxy‐tamoxifen (Z‐4OHtam), with treatment outcomes in an independent cohort of BC patients. Among all patients, metabolite levels did not affect survival. However, in the premenopausal subgroup receiving tamoxifen alone (n = 191) we confirmed an inferior BC ‐specific survival in patients with the previously described serum concentration threshold of Z‐4OHtam ≤ 3.26 nm (HR = 2.37, 95% CI = 1.02–5.48, P = 0.039). The 'dose–response' survival trend in patients categorized to ordinal concentration cut‐points of Z‐4OHtamoxifen (≤ 3.26, 3.27–8.13, > 8.13 nm) was also replicated (P‐trend log‐rank = 0.048). Z‐endoxifen was not associated with outcome. This is the first study to confirm the association between a published active tamoxifen metabolite threshold and BC outcome in an independent patient cohort. Premenopausal patients receiving 5‐year of tamoxifen alone may benefit from therapeutic drug monitoring to ensure tamoxifen effectiveness. [ABSTRACT FROM AUTHOR]

Introduction. Over-treatment of low-risk early breast cancer patients with adjuvant systemic therapies is an important clinical challenge. Better techniques are required which can be used to distinguish between the large group of patients with no residual disease after surgery and consequently no benefit of adjuvant treatment, from the smaller group with high relapse risk. A better integration of available prognostic factors might contribute to improved prediction of clinical outcome. Material and methods. The current study included 346 unselected pTlpNO patients who did not receive adjuvant systemic treatment. In Norway, no patients with this stage were recommended systemic treatment at the time of the study (1995-1998). Histological type, tumour size, grade, vascular invasion (VI), hormone receptor (HR) status, HER2 and Ki67 (cutoff 10%) were analysed. Median follow-up was 86 months for relapse and 101 months for death. Results. Thirty-eight patients experienced relapse, 31 with distant metastasis. Twenty-one patients died of breast cancer. In univariate analysis grade, HER2, HR, VI and Ki67 had impact on clinical outcome (p<0.005, log rank). In multivariate analysis, only grade 1-2 vs. grade 3, HER2, VI, and Ki67 status were significant for disease free survival, distant disease free survival, and/or breast cancer specific survival. These factors were used in combination, to separate patients into groups based on the number of unfavourable factors present [combined prognostic score (CPS) 0-4]. Close to 2/3 of the patients (61.4%) had no unfavourable factor (CPS0), whilst 18.4% had CPS ⩾ 2. Only 3.6% of those with CPS0 developed metastasis (p < 0.001). The outcome was clearly worse for patients with CPS ⩾ 2 (p < 0.001), systemic relapse was detected in approximately 40%. Conclusions. This study indicates that the combined use of grade, VI, HER2 and Ki67 identifies a subgroup of breast cancer patients with a relapse risk that may question the benefit of adjuvant systemic therapy. [ABSTRACT FROM AUTHOR]

The clinical relevance of isolated tumor cell (ITC: ≤0.2 mm) and micrometastasis (MM: >0.2–2.0 mm) in axillary lymph nodes (ALNs) remains unknown. The aim of this study was to determine their prognostic significance. A total of 295 patients considered as pN0 after routine histological assessment, were reevaluated with ten-level cytokeratin immunohistochemistry (IHC) and two-level hematoxylin-eosin sections. Survival rates, i.e. disease-free survival (DFS), distant disease-free survival (DDFS) and breast cancer specific survival (BCSS) were compared with those of reevaluated node-negative patients. A total of 84 patients (28%) had ITC/MM identified on IHC sections. ITC had no impact on survival at a median 8.2 years of follow-up, whereas MM showed a trend toward poorer DFS ( P = 0.091, log rank) and DDFS ( P = 0.066) and significantly reduced BCSS ( P = 0.016). In multivariate analyses, detection of MM was an independent prognostic factor for DDFS ( P = 0.025) and BCSS ( P = 0.01) in adjuvant un-treated patients. Micrometastases (MMs) in axillary lymph nodes have prognostic impact. This was not found for ITC. This finding supports the use of systemic adjuvant therapy in patients with MM. [ABSTRACT FROM AUTHOR]

Representative tumour sections from 468 patients with invasive breast cancer were immunostained for cyclooxygenase-2 (COX-2) and evaluated. The relationships between COX-2 expression, clinical outcome and various clinicopathological variables, including tumour vascularity and disseminated tumour cells (DTC) in the bone marrow were examined. COX-2 expression in invasive breast carcinoma cells was positively associated with oestrogen receptor and/or progesterone receptor positivity (p<0.001). Triple-negative tumours showed no/low COX-2 expression more frequently than other tumour types (p<0.001). Expression of COX-2 was not associated with breast cancer-specific survival (p=0.49, log-rank) or distant disease-free survival (p=0.67, log-rank) for all patients, including lymph node-negative, untreated patients (p>0.14, log-rank). There was also no significant association between COX-2 expression and histological grade, tumour size, nodal status, DTC in bone marrow, p53, HER2, or tumour vascularity. In conclusion, COX-2 expression in this series was associated with the presence of hormone receptors. Low COX-2 expression was observed in triple-negative breast carcinomas.

Manual counting of microvessels as intratumoral microvessel density (MVD) and Chalkley counting have been used in several studies to assess the prognostic impact of vascularity in invasive breast carcinomas. In our present study, the aim was to evaluate the prognostic value of angiogenesis in invasive breast carcinoma assessed by MVD and Chalkley techniques in the same series of patients. A total of 498 breast carcinoma patients with median follow up time 85 months were evaluated. The tumour vascularity was quantified by both manual microvessel count (MVD) and Chalkley count in CD34 stained breast carcinoma slides by a single investigator blinded to clinical information. Other relevant clinicopathological parameters were noted, including breast cancer related death and both loco-regional and systemic relapse. The patients were stratified by converting MVD and Chalkley counts to categorical variables to assess prognostic impact, and results were compared. High vascular grades using MVD count did not demonstrate any prognostic significance for breast cancer specific survival (BCSS) or distant disease free survival (DDFS) either in whole patient group (BCSS, p=0.517, DDFS, p=0.301) or in non-treated node negative patients (p>0.05). Chalkley count showed prognostic significance for both DDFS and BCSS in whole patient group (p<0.001) and also in untreated node negative patient group (p<0.05). In multivariate analysis, Chalkley count, but not MVD, retained the prognostic value for BCSS (p=0.007) and DDFS (p=0.014). The Chalkley count for assessing angiogenesis in invasive breast carcinomas demonstrated prognostic value. The Chalkley method appears to be the better method in estimating the prognostic impact of vascularity in invasive breast carcinomas.