Cecilie Schirmer

Comparison of the clinical significance of occult tumor cells in blood and bone marrow in breast cancer

Immunocytochemical (ICC) detection of disseminated tumor cells (DTC) in bone marrow (BM) in early breast cancer is an independent prognostic factor. The significance of circulating tumor cells (CTC) in peripheral blood (PB) needs further exploration and comparison to DTC detection. PB and BM were prospectively collected from 341 breast cancer patients median 40 months after operation. PB samples were analyzed for tumor cells by a negative immunomagnetic technique (10 × 106 cells/test). BM aspirates were analyzed by standard ICC (2 × 106 cells/test). CTC were present in 10% of the patients and DTC in 14%. Thirty‐seven relapses and 14 breast cancer deaths have occurred at median 66 months after diagnosis. Both CTC‐status and DTC‐status were significantly associated with disease free survival (DFS) (event rate: CTC‐positive 26.5% vs. CTC‐negative 9.1%; DTC‐positive 29.2% vs. DTC‐negative 7.8%) (p < 0.001/p < 0.001, log rank) and breast cancer specific survival (event rate: CTC‐positive 17.6% vs. CTC‐negative 2.6%; DTC‐positive 12.5% vs. DTC‐negative 2.7%) (p < 0.001/p < 0.001). The presence of both CTC and DTC (n = 8) resulted in an especially poor prognosis (p < 0.001). In node negative patients, DTC‐status, but not CTC‐status, predicted differences in DFS (p = 0.006 vs. p = 0.503). Excluding 23 patients with breast cancer‐related events prior to the sample collections, CTC detection was not significantly associated with DFS/distant‐DFS (p = 0.158/0.193), in contrast to DTC detection (p < 0.001/<0.001). Presence of CTC and absence of DTC did not affect DFS (p = 0.516). Applied to early stage disease, CTC analysis of increased volumes of PB appears less sensitive and prognostic than standard DTC analysis. Currently, this does not support an exchange of BM with PB for analysis of occult tumor cells.

Clinical Outcome With Correlation to Disseminated Tumor Cell (DTC) Status After DTC-Guided Secondary Adjuvant Treatment With Docetaxel in Early Breast Cancer

PURPOSE The presence of disseminated tumor cells (DTCs) in bone marrow (BM) predicts survival in early breast cancer. This study explores the use of DTCs for identification of patients insufficiently treated with adjuvant therapy so they can be offered secondary adjuvant treatment and the subsequent surrogate marker potential of DTCs for outcome determination. PATIENTS AND METHODS Patients with early breast cancer who had completed six cycles of adjuvant fluorouracil, epirubicin, and cyclophosphamide (FEC) chemotherapy underwent BM aspiration 2 to 3 months (BM1) and 8 to 9 months (BM2) after FEC. Presence of DTCs in BM was determined by immunocytochemistry using pan-cytokeratin monoclonal antibodies. If one or more DTCs were present at BM2, six cycles of docetaxel (100 mg/m(2), once every 3 weeks) were administered, followed by DTC analysis 1 and 13 months after the last docetaxel infusion (after treatment). Cox regression analysis was used to evaluate disease-free interval (DFI). RESULTS Of 1,066 patients with a DTC result at BM2 and available follow-up information (median follow-up, 71.9 months from the time of BM2), 7.2% were DTC positive. Of 72 docetaxel-treated patients analyzed for DTCs after treatment, 15 (20.8%) had persistent DTCs. Patients with remaining DTCs had markedly reduced DFI (46.7% experienced relapse) compared with patients with no DTCs after treatment (adjusted hazard ratio, 7.58; 95% CI, 2.3 to 24.7). The docetaxel-treated patients with no DTCs after treatment had comparable DFI (8.8% experienced relapse) compared with those with no DTCs both at BM1 and BM2 (12.7% experienced relapse; P = .377, log-rank test). CONCLUSION DTC status identifies high-risk patients after FEC chemotherapy, and DTC monitoring status after secondary treatment with docetaxel correlated strongly with survival. This emphasizes the potential for DTC analysis as a surrogate marker for adjuvant treatment effect in breast cancer.

Disseminated tumor cells as selection marker and monitoring tool for secondary adjuvant treatment in early breast cancer. Descriptive results from an intervention study

BackgroundPresence 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.MethodsA 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.ResultsClinical 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).ConclusionsAfter 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 registrationClin Trials Gov NCT00248703

Disseminated tumor cells in bone marrow following definitive radiotherapy for intermediate or high-risk prostate cancer

The purpose of this study was to explore the prevalence of disseminated tumor cells (DTCs) in bone marrow (BM) of clinically progression‐free prostate cancer (PC) patients at least 2 years after curatively intended radiotherapy (RT) with or without adjuvant hormone treatment.

Abstract P1-08-21: Detection and monitoring of circulating endothelial cells, circulating tumor cells and disseminated tumor cells during neoadjuvant breast cancer treatment including bevacizumab

Background: Results from studies with bevacizumab in addition to traditional neoadjuvant therapy (NAT) indicate a need for predictive biomarkers. As a sub-study of the NeoAva study (a neoadjuvant study), the aim was to investigate the potential association between the presence of circulating endothelial cells (CECs) in peripheral blood (PB), circulating tumor cells (CTCs) in PB and disseminated tumor cells (DTCs) in bone marrow (BM) at different time points during NAT +/- bevacizumab and at one year follow-up, and therapy response. Patients and methods: A total of 150 HER2-negative patients with cT2-4 (≥ 2.5cm) N0-3 M0 status were randomized to receive NAT with or without bevacizumab. In this sub-study, 90 patients have so far been analyzed. Of these, 82 received chemotherapy (FEC100→taxane) and 8 received endocrine therapy (letrozole) + / - bevacizumab. Therapy response was evaluated according to the RECIST criteria and achievement of pathological complete response (pCR). Number of CECs, CTCs and DTCs were assessed at baseline after 12 weeks, at surgery (after 24 weeks) and at one year follow-up. Blood samples were analyzed by CellSearch® to assess CEC and CTC counts. The detection of DTCs was performed by immunocytochemical analysis of 2 × 106 BM mononuclear cells. Results: The pathological complete response rate was 10 out of 90 (11.1%), eight of these patients received bevacizumab. For bevacizumab-treated patients with a change in CEC counts from baseline to time of surgery below median change (27 CECs), 35% (6/17) achieved pCR compared to 6% (1/18) in the group with a CEC count-increase above median change (p = 0.035). The corresponding pCR rates for patients not receiving bevacizumab (median CEC change 131 CECs) were 0% (0/15) and 13% (2/14), respectively. Stepwise testing of thresholds for CEC changes in the bevacizumab-arm revealed significant associations to pCR for change-values between 20 and 40. CTC- and DTC-status or -changes were not associated with tumor response or CEC changes. Conclusion: The presented results indicate that the level of change in the number of circulating endothelial cells during neoadjuvant therapy including bevacizumab is associated with the pathological complete response rate in breast cancer patients. This supports additional testing of CECs as a surrogate marker for response to this treatment. The analyses will be up-dated with results from the rest of the included patients. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P1-08-21.

Abstract P3-06-17: Detection and monitoring of circulating endothelial cells, circulating tumor cells and disseminated tumor cells during neoadjuvant treatment including an anti-angiogenic agent

Withdrawn by Author Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P3-06-17.

P4-06-06: Morphological Categories of ICC-Detected CK+ Cells in Bone Marrow Have Different Prognostic Impact in Breast Cancer.

Introduction: Morphological evaluation increases the specificity of immunocytochemical analysis (ICC) of disseminated tumor cells (DTC) in bone marrow (BM). In the Oslo1 cohort of primary breast cancer (BC) patients (pts), ICC-positive cells detected in BM were prospectively classified into 4 morphological groups: Tumor cells (TC) and un-interpretable cells (UIC i.e. degenerated/fragmented cells), both significantly associated with reduced survival; and hematopoietic cells (HC) and questionable HC (Q-HC), not affecting outcome (Borgen 1999, Naume 2004). The present study revisits these categories, comparing their significance according to therapy and tumor subtype. Materials and methods: Mononuclear cells (MNC) from BM collected at surgery of 761 pts were analyzed by standard ICC, including 2×106 BM MNC stained for cytokeratin by AE1AE3 monoclonal antibodies, and corresponding isotype specific negative controls (neg co; 2×106 BM MNC for most). Cells were classified as TC, UIC, Q-HC or HC. Primary tumor analyses included pTpN stage, grade, hormone receptor (HR) and HER2. Patients were followed for median 99 months. Survival analyses were performed by Kaplan Meyer and Cox regression analysis. Results: Frequency of TC in specific test (AE1AE3) was 16.9%, UIC 14.0%, Q-HC 23.4% and HC 27.2%. By analyzing the entire cohort and numerically correcting for similar cells in neg co, only cells classified as TC or UIC predicted systemic relapse. Analysis of no adjuvant treated (NO-ADJ; n=390) and adjuvant treated (ADJ; n=361) pts separately, revealed that in NO-ADJ pts only those harboring Q-HC experienced reduced survival (DDFS: HR 2.2, 95% CI 1.2−4.1, p=0.01; BCSS: HR 2.7, 95% CI 1.3−5.5, p=0.006). InADJ pts, only TC and UIC predicted relapse (HR 1.8, 95% CI 1.2−2.9, p=0.004) and death (HR 2.0, 95% CI 1.3−3.1, p=0.003). In multivariate analysis for NO-ADJ pts, presence of Q-HC by AE1AE3 remained significant (DDFS: HR 2.1, 95% CI 1.1−4.1, p=0.031; BCSS: HR 2.4, 95% CI 1.1−5.1, p=0.024). By subgroup analysis according to HR and HER2 (where available), prognostic impact of TC and UIC were observed only in HR+/HER2− subgroup (n=538; DDFS:HR 1.9, 95% CI 1.2−3.2, p=0.006; BCSS: HR 3.3, 95% CI 1.9−5.6, p>0.001), and not in HER2+ (n=80), or triple negative pts (n=117). In contrast, presence of Q-HC was associated with worse outcome in triple negative pts (n=115; DDFS:HR 2.9, 95% CI 1.4−6.1, p=0.005; BCSS: HR 2.4, 95% CI 1.1−5.2, p=0.026), with no significant prognostic impact in the other two pts groups. Interestingly, pts habouring ≥ 3 HC in AE1AE3 and in neg co together (8.5% of pts), had improved DDFS (HR 0.60, 95% CI 0.41−0.88, p=0.008) and BCSS (HR 0.57, CI .036-0.91, p=0.018) compared to those with fewer or no HC (p=0.029). Only 4.7% of pts with ≥ 3 HC experienced metastasis, versus 20.8% of the other. Conclusion: Morphological DTC subgroups may differ in clinical significance according to biology/primary tumor subtype and therapy status. This emphasizes the importance of separate analyses of DTC categories and further DTC characterization. False positive cells, probably in the B-cell lineage, might signify an immune-related favorable clinical outcome. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P4-06-06.