Monitoring Of Metastatic Breast Cancer Research Articles

Abstract PO5-15-02: Integration of synonymous mutations and variants of unknown significance for basal and longitudinal characterization of metastatic breast cancer by circulating tumor DNA

Abstract Background: Despite how commonly synonymous mutations (SYN) and unknown significance (VUS) are detected in tissue and circulating tumor DNA (ctDNA) next-generation sequencing (NGS), the potential for these somatic changes to impact personalized approaches in metastatic breast cancer (MBC) is unclear. Emerging evidence suggests that SYN mutations may play a role in cancer biology through changes in mRNA stability, splicing, and gene expression. The aim of this study was to characterize SYN and VUS in a large multicenter consortium and to analyze their interplay with pathogenic variants (PATH). Methods: The study retrospectively analyzed a multi-institutional cohort comprising 1189 patients with MBC characterized for ctDNA using NGS (Guardant360™, Guardant Health) before treatment start (BL) and at disease progression (PD). Pathway classification was defined based on prior research (Sanchez-Vega F et al, Cell. 2018). Single Nucleotide Variations (SNVs) were annotated for oncogenicity (OncoKB) and protein domain (UniProtKB). Associations between clinical characteristics and pathway classifications for SYN and VUS were explored by multinomial logistic regression; survival was tested through Cox regression in terms of overall survival (OS). Results: Hormone-receptor positive (HR+)/HER2 negative (luminal-like) was the most represented subtype (68.7%) followed by HER2-positive (14.6%), and triple-negative (TNBC) (16.7%). PATH were mainly detected in the TP53 DNA binding domain, ESR1 ligand binding domain, helical and catalytic domains of PIK3CA, SYN in the ligand binding domain of ESR1, MET Sema, KIT protein kinase domain, and FGFR1 Ig-like C2-type 1. In contrast, VUS were mainly found in the ESR1 ligand binding domain, MET Sema, ATM PI3K/PI4K catalytic domain, DDR2 Protein kinase and PIK3CA C2 PI3K-type. Analyzing pathways, after multivariable multinomial logistic regression, RAS SYN, PI3K and ESR PATH were significantly associated with HER2+ MBC (respectively RRR=4.14, P=0.046; RRR = 0.474, P < 0.001 and RRR = 0.306, P < 0.001). TNBC was associated with PATH PI3K (RRR = 0.366, P < 0.001), PATH P53 (RRR = 4.71, P < 0.001) and PATH RAS (RRR = 0.362, P = 0.026, 95% CI 0.147 - 0.888). In luminal-like MBC, a significant impact on OS was observed for SYN P53 (HR = 2.53, P = 0.008) and PATH ER (HR = 1.54, P < 0.001), PATH P53 (HR = 1.56, P < 0.001), PATH cell-cycle (HR = 1.64, P = 0.0299), and PATH RAS (HR = 1.62, P = 0.010). An impact on OS was observed in TNBC for SYN cell-cycle (HR = 5.41, P = 0.006), VUS ESR (HR = 2.87, P = 0.018), PATH P53 (HR = 1.84, P = 0.012) and PATH cell-cycle (HR = 5.07, P < 0.001). VUS MYC (HR = 38.5, P = 0.001) and PATH PI3K3 (HR = 2.32, P = 0.022) were significant among HER2 positive. Differences in SYN, VUS and PATH alterations were then compared between baseline and progressive disease. While a significant increase in mutant allele frequency (MAF) was observed for PATH (P=0.0010) and VUS (P=0.0152) at PD, no differences were highlighted for SYN (P=0.8362). Conclusions: In our cohort we characterized the MBC for PATH, VUS and SYN utilizing the ctDNA. In this study, considering the complex genomic landscape of MBC, we demonstrated that both SYN and VUS mutations had an impact on the survival outcome. Therefore, further analysis to validate the prognostic and predictive role of SYN and VUS, with or without PATH mutations, are needed to better characterize and monitoring of MBC through ctDNA. Moreover, functional studies are needed to better understand their role in the nuanced biology of MBC. Citation Format: Elisabetta Molteni, Carolina Reduzzi, Andrew Davis, Lorenzo Foffano, Arielle Medford, Katherine Clifton, Whitney L. Hensing, Marko Velimirovic, Ami N. Shah, Laura Munoz Arcos, Charles S. Dai, Jennifer C. Keenan, Elyssa Denault, William Gradishar, Giuseppe Damante, Amir Behdad, Lorenzo Gerratana, Fabio Puglisi, Cynthia Ma, Aditya Bardia, Massimo Cristofanilli. Integration of synonymous mutations and variants of unknown significance for basal and longitudinal characterization of metastatic breast cancer by circulating tumor DNA [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO5-15-02.

2-[18F]FDG-PET/CT is a better predictor of survival than conventional CT: a prospective study of response monitoring in metastatic breast cancer

This study aimed to compare CE-CT and 2-[18F]FDG-PET/CT for response monitoring metastatic breast cancer (MBC). The primary objective was to predict progression-free and disease-specific survival for responders vs. non-responders on CE-CT and 2-[18F]FDG-PET/CT. The secondary objective was to assess agreement between response categorization for the two modalities. Treatment response in women with MBC was monitored prospectively by simultaneous CE-CT and 2-[18F]FDG-PET/CT, allowing participants to serve as their own controls. The standardized response evaluation criteria in solid tumors (RECIST 1.1) and PET response criteria in solid tumors (PERCIST) were used for response categorization. For prediction of progression-free and disease-specific survival, treatment response was dichotomized into responders (partial and complete response) and non-responders (stable and progressive disease) at the first follow-up scan. Progression-free survival was defined as the time from baseline until disease progression or death from any cause. Disease-specific survival was defined as the time from baseline until breast cancer-specific death. Agreement between response categorization for both modalities was analyzed for all response categories and responders vs. non-responders. At the first follow-up, tumor response was reported more often by 2-[18F]FDG-PET/CT than CE-CT, with only fair agreement on response categorization between the two modalities (weighted Kappa 0.28). Two-year progression-free survival for responders vs. non-responders by CE-CT was 54.2% vs. 46.0%, compared with 59.1% vs. 14.3% by 2-[18F]FDG-PET/CT. Correspondingly, 2-year disease-specific survival were 83.3% vs. 77.8% for CE-CT and 84.6% vs. 61.9% for 2-[18F]FDG-PET/CT. Tumor response on 2-[18F]FDG-PET/CT was significantly associated with progression-free (HR: 3.49, P < 0.001) and disease-specific survival (HR 2.35, P = 0.008), while no association was found for tumor response on CE-CT. In conclusion, 2-[18F]FDG-PET/CT appears a better predictor of progression-free and disease-specific survival than CE-CT when used to monitor metastatic breast cancer. In addition, we found low concordance between response categorization between the two modalities.Trial registration: Clinical.Trials.gov. NCT03358589. Registered 30/11/2017-Retrospectively registered, http://www.ClinicalTrials.gov.

Abstract 3433: Global proteomic analysis of urine using a label-free LC-MS/MS approach identifies potential biomarkers of metastatic breast cancer

Abstract Introduction: Breast cancer (BC) is a complex heterogenous disease and is a leading cause of death in women. Following primary BC therapy ~40% of patients will develop metastatic BC (MBC) and 60-70% of the recurrent disease will occur at distant sites. Median survival of MBC patients with distant recurrence is ~2-3 yr. While the early detection and monitoring of BC progression are essential to improve prognosis and reduce BC-related mortality, there is a lack of surveillance strategies for monitoring patients for distant recurrence of MBC. The aim of our study was to identify urinary biomarkers for detection and monitoring of MBC. Method: We have conducted a comparative label-free LC-MS/MS analysis of the urinary proteome of patients with MBC and healthy age-matched controls (HC). A hybrid quadrupole time of flight (Q-Tof™) mass spectrometer was used for urine analysis via liquid chromatography (LC) with tandem mass spectrometry (MS/MS). Three different database search algorithms were used for peptide identification including MASCOT, Protein Lynx Global Server and Ion Accounting software. Only peptides detected in at least two out of three replicated LC-MS experiments were counted towards protein identification. Identified candidate biomarkers were validated via ELISA assays. Results: Using this approach, we identified ~400 urinary proteins of which ~104 and 89 were unique to the MBC and HC groups, respectively. Upregulated proteins in the MBC cohort were associated with angiogenesis, apoptosis, proteolysis, ECM regulation, and cell adhesion pathways. Using bioinformatic and in silico approaches, we identified a specific metastasis signature comprised of CALB1, S100A8, ZAG, VTN and TN. Diagnostic accuracies of these candidate markers were validated using independent training and validation sets according to the REMARK criteria. Urinary VTN (uVTN) and uTN levels correlated significantly with disease status in MBC patients when compared to patients with locally invasive BC and HC. Interestingly, a small subset of patients with bone metastasis also had significantly higher uVTN levels. A multiplexed marker panel of uVTN and uTN could discriminate between HC and MBC groups (AUC=0.831, P&amp;lt;0.001). Blinded testing indicated that uVTN levels (cutoff&amp;gt;1018 ng/ml) could discriminate (AUC=0.782, P&amp;lt;0.006) between MBC and HC. Longitudinal analysis of samples from MBC patients indicated a strong correlation between uVTN levels and disease status. Conclusions: Our findings suggest that uVTN and uTN have the potential to serve as biomarkers for the detection of MBC. While validation in larger cohorts is necessary, these results may be useful in the development of noninvasive diagnostic tests for monitoring BC progression and recurrence. [Supported by: the Advanced Medical Research Foundation, the Jo Ann Webb Fund for Angiogenesis Research and the S. Elizabeth O’Brien Trust] Citation Format: Roopali Roy, Elisa Schunkert, Petra Olivova, Martin Gilar, Scott Geromanos, Guo-Zhong Li, John Gebler, Adelle Dagher, Andrew El-Hayek, Rama Aldakhlallah, David Zurakowski, Susan Pories, Marsha A. Moses. Global proteomic analysis of urine using a label-free LC-MS/MS approach identifies potential biomarkers of metastatic breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3433.

Clinical Impact of FDG-PET/CT Compared with CE-CT in Response Monitoring of Metastatic Breast Cancer.

Simple SummaryThe method of treatment evaluation in patients with chronic breast cancer may affect clinical decision making and treatment protocols. In this study, we compared the two imaging modalities for the evaluation of treatment responses in 65 patients with spread breast cancer. We included 34 patients who underwent contrast-enhanced computed tomography (CE-CT) and 31 patients who underwent positron emission tomography/computed tomography (FDG-PET/CT). Then, we compared the response categories and clinical effects within the two modalities during a follow-up period of an average of 17.3 months. Our results showed that CE-CT modality reported more scans as stable disease, while FDG-PET/CT modality reported regressive metastatic disease more often. This means that FDG-PET/CT responds more precisely with respect to the changes in patients’ clinical condition, while CE-CT tends to report stable disease in most of the scans. Therefore, FDG-PET/CT may be a more suitable imaging modality than CE-CT for the evaluation of treatment in patients with metastatic breast cancer.We compared response categories and impacts on treatment decisions for metastatic breast cancer (MBC) patients that are response-monitored with contrast-enhanced computed-tomography (CE-CT) or fluorodeoxyglucose-positron emission tomography/computed tomography (FDG-PET/CT). A comparative diagnostic study was performed on MBC patients undergoing response monitoring by CE-CT (n = 34) or FDG-PET/CT (n = 31) at the Odense University Hospital (Denmark). The responses were assessed visually and allocated into categories of complete response (CR/CMR), partial response (PR/PMR), stable disease (SD/SMD), and progressive disease (PD/PMD). Response categories, clinical impact, and positive predictive values (PPV) were compared for follow-up scans. A total of 286 CE-CT and 189 FDG-PET/CT response monitoring scans were performed. Response categories were distributed into CR (3.8%), PR (8.4%), SD (70.6%), PD (15%), and others (2.1%) by CE-CT and into CMR (22.2%), PMR (23.8%), SMD (31.2%), PMD (18.5%), and others (4.4%) by FDG-PET/CT, revealing a significant difference between the groups (P < 0.001). PD and PMD caused changes of treatment in 79.1% and 60%, respectively (P = 0.083). PPV for CE-CT and FDG-PET/CT was 0.85 (95% CI: 0.72–0.97) and 0.70 (95% CI: 0.53–0.87), respectively (P = 0.17). FDG-PET/CT indicated regression of disease more frequently than CE-CT, while CE-CT indicated stable disease more often. FDG-PET/CT seems to be more sensitive than CE-CT for monitoring response in metastatic breast cancer.

Interrater Agreement and Reliability of PERCIST and Visual Assessment When Using 18F-FDG-PET/CT for Response Monitoring of Metastatic Breast Cancer.

Response evaluation at regular intervals is indicated for treatment of metastatic breast cancer (MBC). FDG-PET/CT has the potential to monitor treatment response accurately. Our purpose was to: (a) compare the interrater agreement and reliability of the semi-quantitative PERCIST criteria to qualitative visual assessment in response evaluation of MBC and (b) investigate the intrarater agreement when comparing visual assessment of each rater to their respective PERCIST assessment. We performed a retrospective study on FDG-PET/CT in women who received treatment for MBC. Three specialists in nuclear medicine categorized response evaluation by qualitative assessment and standardized one-lesion PERCIST assessment. The scans were categorized into complete metabolic response, partial metabolic response, stable metabolic disease, and progressive metabolic disease. 37 patients with 179 scans were included. Visual assessment categorization yielded moderate agreement with an overall proportion of agreement (PoA) between raters of 0.52 (95% CI 0.44–0.66) and a Fleiss kappa estimate of 0.54 (95% CI 0.46–0.62). PERCIST response categorization yielded substantial agreement with an overall PoA of 0.65 (95% CI 0.57–0.73) and a Fleiss kappa estimate of 0.68 (95% CI 0.60–0.75). The difference in PoA between overall estimates for PERCIST and visual assessment was 0.13 (95% CI 0.06–0.21; p = 0.001), that of kappa was 0.14 (95% CI 0.06–0.21; p < 0.001). The overall intrarater PoA was 0.80 (95% CI 0.75–0.84) with substantial agreement by a Fleiss kappa of 0.74 (95% CI 0.69–0.79). Semi-quantitative PERCIST assessment achieved significantly higher level of overall agreement and reliability compared with qualitative assessment among three raters. The achieved high levels of intrarater agreement indicated no obvious conflicting elements between the two methods. PERCIST assessment may, therefore, give more consistent interpretations between raters when using FDG-PET/CT for response evaluation in MBC.

Abstract 4416: Plasma thymidine kinase activity in patients with luminal metastatic breast cancer treated with Palbociclib within the phase II TREnd trial

Abstract Background: The CDK4/6 inhibitor palbociclib (P) is approved for the treatment of luminal metastatic breast cancer (MBC) in combination with endocrine therapy (ET). It leads to reduced phosphorylation of the Rb protein resulting in a decrease in E2F activity and eventual cell cycle arrest. Thymidine kinase 1 is a well-known cancer proliferation marker downstream of the E2F pathway, whose activity can be measured in plasma samples as readout of tumor proliferation. Circulating thymidine kinase activity (TKa) is a prognostic marker in MBC patients (pts) treated with ET, both when measured at baseline and during treatment. TKa has been previously shown to decrease in pts treated with neoadjuvant P+ET for 15 days, which was attributed as pharmacodynamic change on P treatment. The predictive value of TKa changes during treatment with P, as well as the dynamics of TKa changes on P-containing treatments are not yet comprehensively defined. Here we investigate the role of plasma TKa measured at different timepoints in a cohort from the TREnd study (NCT02549430). Methods: TREnd was a randomized phase II trial that allocated 115 pts with moderately pre-treated luminal MBC to receive single-agent P or P plus the same ET agent that was received in the prior line of therapy. Plasma samples were collected at baseline (T0; n=45), at day 1 of cycle 2 (T1; n=45) and at disease progression (T2; n= 36) from 46 consenting pts. TKa was measured with DiviTum®, a refined ELISA-based assay. Patients were dichotomized as high/low at T0 based on an optimal cut-off (260 Du/L) determined by maximally selected rank statistics, and on the median value at T2 (250 Du/L). Dynamic changes between T0 and T1 were deemed meaningful if &amp;gt;10% of T1 or T0, whichever was greatest. Clinical outcome was estimated using the Kaplan-Meier method. Results: Median TKa (mTKa) at T0 was 73 Du/L (range 20-4302). As expected, P-containing treatment reduced mTKa levels at T1 (37 Du/L, range 20-4504). Conversely, at disease progression, TKa increased compared to T0 (mTKa at T2, 250 Du/L, range 20-3653). Median time to progression (mTTP) in pts with low TKa at T0 (n= 33) was 8.5 months, compared to 5.6 months in pts with high TKa (n= 12). Interestingly, pts with an increase in TKa at T1 (n=9) had a mTTP of only 3.1 months compared to pts with stable/reduced TKa (N=35), who showed a mTTP of 9 months. Considering the potential significance of TKa measured at disease progression, pts with high levels at T2 (n=18) had a worse outcome on subsequent post-study treatment (both chemotherapy and ET) compared to those with lower levels (n=18) (mTTP at T2, 2.9 vs 8.9 months, respectively). Conclusions: These data suggest for the first time that TKa may be a useful prognostic biomarker for non-invasive monitoring of MBC in the context of treatment with P. These results warrant further investigation in larger sample sets. Citation Format: Luca Malorni, Chiara Biagioni, Francesca De Luca, Martina Bonechi, Giuseppe Curigliano, Alessandro M. Minisini, Erica Moretti, Mattias Bergqvist, Karin Mattsson, Emanuela Risi, Ilenia Migliaccio, Stefania Vitale, Stefano Gabellini, Amelia McCartney, Irene De Santo, Francesca Galardi, Giulia Boccalini, Matteo Benelli, Lorenzo Rossi, Laura Biganzoli, Angelo Di Leo. Plasma thymidine kinase activity in patients with luminal metastatic breast cancer treated with Palbociclib within the phase II TREnd trial [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4416.

Controversies in monitoring metastatic breast cancer during systemic treatment. Results of a GIM (Gruppo Italiano Mammella) survey

BackgroundThe optimal strategy for monitoring metastatic breast cancer (M-MBC) is unclear. Nevertheless, M-MBC influences patient's quality of life and it affects the use of resources in terms of both drugs and diagnostic exam prescription. We aim to disclose oncologists' approach on M-MBC, identifying controversial areas. MethodsAn anonymous online survey was conducted among GIM members who, based on their on-field experience, shared their own method for M-MBC planning. Chi-square tests and Fisher exact tests were used as appropriate. ResultsThe survey was completed by 256 recipients (51%). The majority of them were medical oncologists. Approximately 50% of respondents reported that M-MBC was primarily based on the monitoring strategies used in clinical trials, and for 70% of them M-MBC should be evidence-based. Areas of controversies included the primary goal of M-MBC, the use of tumour markers, the optimal timing for baseline assessment and frequency of repeating testing. Respondents agreed on planning M-MBC before treatment's start and on discussing with the patient about the M-MBC strategy and on choosing CT-scan as the preferred reassessment imaging method. The most relevant factors influencing the M-MBC strategy were performance status, triple negative histology, exam's contraindication, the presence of clinically measurable disease, and treatment safety profile; on the contrary, patients' socio-economic status, exam's cost, and hospital's logistic limits were less relevant. Experienced oncologists seemed less prone to intensive follow-up. ConclusionM-MBC strategy still has controversial issues and its potential clinical effects for breast cancer patients need ad hoc studies.

Monitoring metastatic breast cancer (M-MBC) during treatment: a GIM (Gruppo Italiano Mammella) survey

Monitoring metastatic breast cancer (M-MBC) during treatment: a GIM (Gruppo Italiano Mammella) survey

Strategy of monitoring metastatic breast cancer (M-MBC) in clinical practice: more or less intensive?

Strategy of monitoring metastatic breast cancer (M-MBC) in clinical practice: more or less intensive?

Circulating tumor cells (CTCs) and circulating tumor DNA (ctDNA) longitudinal monitoring of metastatic breast cancer (MBC).

e23030Background: Liquid biopsy provides a real-time assessment of MBC. We evaluated enumeration of CTCs and analysis of ctDNA to assess prognosis and detect molecular abnormalities in MBC. Methods...

Estrogen receptor α determination in serum, cell lysates and breast cancer cells using an amperometric magnetoimmunosensing platform

An electrochemical magnetoimmunosensor for the determination of estrogen receptor α (ERα) protein in complex samples (serum and cell lysates) able to discriminate between ERα positive and negative breast cancer cells is reported. Specifically functionalized magnetic microbeads with sandwich immunocomplexes and amperometric detection at disposable screen-printed carbon electrodes (SPCEs) resulted in highly selective and sensitive ERα detection with a detection limit of 19pgmL−1. This magnetoimmunosensing platform was successfully applied to the quantitation of ERα in spiked human serum and cell lysates samples without any matrix effect with an advantageous performance in terms of simplicity and assay times over commercial ELISA assays. The biosensor capability for assessing ERα in intact breast cancer cells makes it competitive with conventional strategies providing rapidly quantitative and reliable results on this relevant biomarker currently used in the clinical practice for diagnosis, follow-up and monitoring of metastatic breast cancer.

Electrochemical Magnetoimmunosensor for Progesterone Receptor Determination. Application to the Simultaneous Detection of Estrogen and Progesterone Breast‐cancer Related Receptors in Raw Cell Lysates.

AbstractAn electrochemical magnetoimmunosensor for the determination of progesterone receptor (PR) making use of functionalized magnetic microbeads (MBs), a sandwich‐type immunoassay and amperometric detection at disposable screen‐printed carbon electrodes (SPCEs) was described for the first time in this work. This selective and sensitive magnetoimmunosensor, with a detection limit of 22 pg mL−1, was successfully applied to the determination of PR in raw cell lysates with just a dilution as sample treatment and exhibiting no matrix effect. The advantageous performance in terms of simplicity and assay time over commercial ELISAs makes it competitive with conventional strategies providing rapidly quantitative and reliable results on this biomarker currently used in the clinical practice for diagnosis, follow‐up and monitoring of metastatic breast cancer. In addition, a dual magnetoimmunosensor using screen‐printed dual carbon electrode (SPdCE) has been implemented for the simultaneous determination of PR and estrogen receptor α (ERα) in cell lysates. This dual receptor determination allows a more accurate diagnosis enabling a more efficient treatment. The usefulness of this dual magnetoimmunosensing platform was demonstrated by discriminating raw lysates of two types of cancer cells (MCF‐7 and MDA‐MB‐436) with significantly different PR/ERα expression levels.

Electrochemical immunosensor for the analysis of the breast cancer biomarker HER2 ECD

Human epidermal growth factor receptor 2 (HER2) is a breast cancer biomarker that plays a major role in promoting breast cancer cell proliferation and malignant growth. The extracellular domain (ECD) of HER2 can be shed into the blood stream and its concentration is measurable in the serum fraction of blood. In this work an electrochemical immunosensor for the analysis of HER2 ECD in human serum samples was developed. To achieve this goal a screen-printed carbon electrode, modified with gold nanoparticles, was used as transducer surface. A sandwich immunoassay, using two monoclonal antibodies, was employed and the detection of the antibody–antigen interaction was performed through the analysis of an enzymatic reaction product by linear sweep voltammetry. Using the optimized experimental conditions the calibration curve (ip vs. log[HER2 ECD]) was established between 15 and 100ng/mL and a limit of detection (LOD) of 4.4ng/mL was achieved. These results indicate that the developed immunosensor could be a promising tool in breast cancer diagnostics, patient follow-up and monitoring of metastatic breast cancer since it allows quantification in a useful concentration range and has an LOD below the established cut-off value (15ng/mL).

Abstract P3-13-11: Unexpected large treatment and outcome variations in metastatic breast cancer: Results of a hormone receptor positive cohort study in the Netherlands

Abstract Introduction Treatment of patients with metastatic breast cancer (MBC) generally has a palliative intent. The systemic treatment approach is based on hormone receptor (HR) and human epidermal growth factor receptor 2 (HER2) status. However, with a variety of available agents and major changes in adjuvant breast cancer treatment, treatment decisions in MBC are complex. The objective of this study was to present treatment patterns of HR positive metastatic breast cancer patients, categorized by HER2 status. Patients and methods All patients diagnosed with HR positive MBC between 2007 and 2009 (irrespective of date of primary diagnosis) in eight participating hospitals in the South-East part of the Netherlands were included and all medical charts were reviewed. Patient and tumor characteristics as well as treatments and outcomes were collected. Survival was estimated using the Kaplan-Meier method. Results A total of 611 patients had HR positive MBC; 529 of these (86.6%) had a negative HER2 status. Median survival after diagnosis of distant metastases was 22.1 months for the HR+/HER2-subtype and 32.9 months for the HR+/HER2+ subtype. In the HR+/HER2- subgroup 7% of patients did not receive any palliative systemic treatment and in the HR+/HER2+ subgroup 8.5% of patients did not receive any palliative systemic treatment. In the HR+/HER2- subgroup, first given palliative systemic treatment was chemotherapy in 22% and endocrine therapy in 71% of patients. Of the patients starting with chemotherapy, 15% had bone metastases only, 37% had visceral metastases and 39% had multiple metastatic sites compared to 49%, 20% and 25% respectively for those starting with endocrine treatment. Of the patients starting with chemotherapy, 38% had progressive disease during or after this line of treatment and 62% had either response or stable disease. In the HR+/HER2+ subgroup, 59.8% of patients received chemotherapy as first palliative systemic treatment (in two thirds of these patients targeted therapy was added to the chemotherapy) and 31.7% received endocrine therapy with or without trastuzumab. Of the patients starting with chemotherapy (+/- targeted therapy) 20% had bone metastases only, 39% visceral metastases and 37% had multiple metastatic sites versus respectively 23%, 31% and 35% for the patients starting with endocrine therapy. Of the patients starting with chemotherapy (+/- targeted therapy), 17% had progressive disease during or after this line of treatment and 83% had either response or stable disease. Conclusion A considerable proportion of HR+ MBC patients received chemotherapy as first given treatment despite HR positive status and presence of bone metastases only or limited visceral metastases, which is more than expected based on current treatment guidelines. Daily-practice monitoring of MBC, as we have reported here, may improve future treatment decision-making. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P3-13-11.

Validity of bone marker measurements for monitoring response to bisphosphonate therapy with zoledronic acid in metastatic breast cancer

Bone is the most common site of metastasis in breast cancer. Detection relies on imaging technology which is costly and can only be performed to a certain degree. Bone markers are non-invasive, inexpensive and may potentially serve as predictive and prognostic surrogate endpoints in detecting bone metastases and response to bisphosphonates. This study analyzed the value of the serum bone turnover markers PINP and ICTP for bone metastases in metastatic breast cancer patients receiving zoledronic acid. The results were compared with the serum levels of CEA and CA 15-3, and analyzed with respect to the number of bone metastases as well as clinical response. Forty patients with confirmed bone metastases who received chemotherapy and/or hormonal therapy and zoledronic acid i.v. q4 weeks participated in the present study. blood (5 ml) was collected at the start of the study and q3 months for a period of one year for the analysis of PINP, ICTP, CEA and CA 15-3 using radioimmunoassays and ELISA, respectively. Imaging of bone metastases was performed at the same time points. In 29 out of 40 patients, more than 3 bone metastases were confirmed by imaging and 11 out of 40 patients presented with 3 or less. At the start of the study, the median value for ICTP was 6 µg/l and for PINP 58.7 µg/l. At the end of the study the median values were 4.5 µg/l for ICTP and 21 µg/l for PINP. When patients were stratified into responders and non-responders, a decrease in both PINP (P<0.0001) and ICTP (P=0.048) was observed for the responders, while the level of ICTP (P=0.02) increased for the non-responders. Serum PINP and ICTP concentrations were significantly different when patients were stratified into groups of those having more than 3 bone metastases and 3 or less, respectively (P<0.05). CEA and CA 15-3 levels did not differ with respect to the number of bone metastases, while the tumor marker levels determined at the end of the study significantly distinguished responders from non-responders (P=0.002 and P=0.004). In conclusion, in contrast to serum tumor markers, the determination of PINP and ICTP allows inferences to the number of bone metastases and appears to be a useful tool for prediction and monitoring metastatic breast cancer patients undergoing bisphosphonate therapy with zoledronic acid for the treatment of bone metastases.

876 Non-invasive Monitoring of Metastatic Breast Cancer by Circulating Tumour DNA – a Comparison With Circulating Tumour Cells

876 Non-invasive Monitoring of Metastatic Breast Cancer by Circulating Tumour DNA – a Comparison With Circulating Tumour Cells

SU-E-J-198: Bioluminescence Monitoring of Metastatic Breast Cancer: Quantitative Assessment of Radiation Treatment Effects and Tracking of Tumor Cells.

To evaluate radiation treatment effects on mammary carcinoma cells, quantitative photon radiance were monitored to track light-emitting cancer cells and metastasis using in vivo bioluminescence imaging. Eight female BALB/c mice aged 8 weeks were orthotopically injected with 5×104/cc 4T1 tumor cells into the abdominal mammary gland. The firefly luciferase-based bioluminescence images were acquired every 2-3 days for 1 month. Bioluminescent intensity was analyzed in average surface radiance (photons/sec/cm2 /sr) taken in 3-dimensional bioluminescence tomography (BLT). After 1 week, single-radiation dose of 20 Gy was delivered by orthovoltage X-rays. Variation of detected bioluminescence signals emitted from molecular cancer cells was depicted on BLT images. To delineate tumor volumes according to bioluminescence intensity on anatomical images for radiation therapy, BLT images were registered with the micro computed tomography (CT) images using surface-constrained warping. Multispectral BLT images elaborated on early detection of cancer cells, characteristics of tumor growth, and metastasis for more accurate determination of internal bioluminescent sources. The radiation-treated mice having only primary tumor volumes showed 67% decrease in bioluminescent signals, while the mice with metastatic cancer cells suggested 88% reduction, as compared to the control group. Registration of BLT with CT images guided molecular cancer cells on anatomical coordinates. The BLT imaging was a useful tool to localize cancer cells and to quantify radiation response. Application of BLT led to more accurate definition of tumor volumes including molecular probe-based microscopic cancer cells. Monitoring of bioluminescence signals enables to diagnose real-time metastatic behavior of cancer cells and determine optimal radiation treatment strategies adapted to tumor characteristics.

P5-13-18: A Study of the Usefulness of Tumor Markers CA 15–3 and TPS in Monitoring of Different Subgroups of Metastatic Breast Cancer.

Abstract Background: The sensitivity and specificity of the tumor markers cancerantigen 15–3 (CA15-3) and the more seldom used tissue polypeptide specific antigen (TPS) for detecting and monitoring metastatic breast cancer are relatively well known. However, most of the studies in this area were performed before the era of breast cancer subtyping and some recent reports have showed differences in the sensitivity of CA 15–3 among different subtypes of breast cancer. This implies that the usefulness of a tumor marker could vary among different subgroups. We performed a retrospective study to find out the usefulness of CA 15–3 and TPS in long-term monitoring of metastatic breast cancer subgrouped by hormonal and HER2−receptor status. Patients and methods: CA 15–3 was routinely assessed regularly in all patients with metastatic breast cancer since 2003, while TPS was assessed during years 2005 — 2009. In total 142 patients (median age 60 years, range 22–87) with metastatic breast cancer were identified in the local data base and included. The total number of analyzed CA 15–3 samples were 4232 (mean 30/patient) and TPS 1993 (mean 14/patient). In each patient the results of the markers were compared with disease response and progression during different lines of therapy and the patients were grouped based on the ratio of correct tumor marker values: “1” (useless) if 0–49% of the values were regarded as true, “2” (poor) if 50–74% were true, “3” (good) if 75–94% were true and finally “4” (excellent) if 95–100% of the values were considered true. If a patient had no elevated values during follow-up, she was reported as tumor marker negative: “0”. Results: In general, the usefulness of CA 15–3 was excellent (4) in 50% of patients and good (3) in 16% more, compared with only 12% and 23% for TPS. Twenty-three percent and 25%, respectively were marker negative (0). There was a marked difference in the different subgroups. CA 15–3 was very useful (group 3 + 4) in estrogen and progesterone receptor positive patients (79%) and hormonal positive HER2 positive patients (82%), compared with only 40% in the triple negative group and 27% of the patients in the hormonal negative HER2 positive group. TPS showed the opposite tendency with less value in hormonal positive patients (only 31% in group 3 and 4) compared with 40% in triple negative and 45% the hormonal negative HER2 positive group. Other factors with strong positive influence of the value of the marker were high absolute values, increased value at diagnosis of metastatic disease (initial value) and visceral metastases. Conclusion: Metastatic breast cancer can be well monitored using CA 15–3 in about 80% of the cases if the tumor is estrogen or progesterone positive. Hormonal receptor negative disease may be well monitored with CA 15–3 and/or TPS if there are elevated initial values, high absolute values and visceral disease. HER2−status seems not to influence the usefulness of CA 15–3 and TPS. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P5-13-18.

A real-time quantitative reverse transcriptase polymerase chain reaction (RT–PCR) to detect breast carcinoma cells in peripheral blood

The detection of occult carcinoma cells in patients with breast cancer has been shown to predict disease recurrence and metastasis. To improve on molecular detection of breast carcinoma cells in blood, we have developed a sensitive and quantitative assay using real-time quantitative RT-PCR identifying transcripts of the cytokeratin-19 (CK19) gene. This real-time quantitative RT-PCR is sensitive, accurate and has a high reproducibility within a wide dynamic range, which permits simultaneous quantitative analysis of samples with varying input concentrations. Furthermore, the procedure offers several technical advantages over classic quantitative PCR methods (competitive RT-PCR, Northern blotting) such as decreased likelihood of contamination due to absence of post-PCR manipulations, high sample throughput because of absence of post-PCR processing time (no agarose gel electrophoresis). In this pilot study, we detected significantly elevated CK19 transcript levels in < 10% of the volunteers, in +/- 30% of stage I-IIIa patients preoperatively and in > 70% of the and stage IV breast cancer patients. Analyses using this real time quantitative RT-PCR for CK19 mRNA may prove to have clinical implications in the assessment of circulating tumour cells in peripheral blood, micrometastases in bone marrow or lymph nodes in breast cancer patients. Application of this technique in a clinical population may improve diagnosis and monitoring of metastatic breast cancer and its validation is currently ongoing.

Description of a Computer Program to Assess Cancer Antigen 15.3, Carcinoembryonic Antigen, and Tissue Polypeptide Antigen Information during Monitoring of Metastatic Breast Cancer

It is time-consuming to process and compare the clinical and marker information registered during monitoring of breast cancer patients. To facilitate the assessment, we developed a computer program for interpreting consecutive measurements. The intraindividual biological variation, the analytical precision profile, the cutoff limit, and the detection limit for each marker are entered and stored in the program. The assessment procedure for marker signals considers the analytical and biological variation of the applied markers. The software package contains a database that can store the interpretation of the measurements as evaluation codes together with patient demographics, information about treatment type, dates for treatment periods, control periods, and evaluation codes for clinical activity of disease. The consecutive concentrations for a patient are imported temporarily into the program from outside sources and presented graphically. Marker concentrations to be compared are selected with the computer mouse and the significance of the difference is calculated by the program. The program has an option for calculating the lead time of marker signals vs clinical information. The program facilitates the monitoring of individual breast cancer patients with tumor marker measurements. It may also be implemented in trials investigating the utility of potential new markers in breast cancer as well as in other malignancies.