Lot A. Devriese

A multimarker QPCR-based platform for the detection of circulating tumour cells in patients with early-stage breast cancer

Background:The detection of circulating tumour cells (CTCs) has been linked with poor prognosis in advanced breast cancer. Relatively few studies have been undertaken to study the clinical relevance of CTCs in early-stage breast cancer.Methods:In a prospective study, we evaluated CTCs in the peripheral blood of 82 early-stage breast cancer patients. Control groups consisted of 16 advanced breast cancer patients and 45 healthy volunteers. The CTC detection was performed using ErbB2/EpCAM immunomagnetic tumour cell enrichment followed by multimarker quantitative PCR (QPCR). The CTC status and common clinicopathological factors were correlated to relapse-free, breast cancer-related and overall survival.Results:Circulating tumour cells were detected in 16 of 82 (20%) patients with early-stage breast cancer and in 13 out of 16 (81%) with advanced breast cancer. The specificity was 100%. The median follow-up time was 51 months (range: 17–60). The CTC positivity in early-stage breast cancer patients resulted in significantly poorer relapse-free survival (log rank test: P=0.003) and was an independent predictor of relapse-free survival (multivariate hazard ratio=5.13, P=0.006, 95% CI: 1.62–16.31).Conclusion:The detection of CTCs in peripheral blood of early-stage breast cancer patients provided prognostic information for relapse-free survival.

Circulating tumor cell detection in advanced non-small cell lung cancer patients by multi-marker QPCR analysis

BACKGROUND The aim of this study was to explore circulating tumor cell (CTC) detection in advanced non-small cell lung cancer (NSCLC). CTCs may not only serve as a prognostic marker in selected tumor types, but may also be useful as pharmacodynamic marker in drug development. METHODS Fourty-six advanced NSCLC patients and fourty-six healthy controls were included in the study and 8.0 ml of peripheral blood was obtained from each of the participants. Immunomagnetic bead enrichment for cells expressing epithelial cell adhesion molecule (EpCAM) was performed, followed by multi-marker quantitative real-time PCR of a panel of marker genes: cytokeratin 7 (CK7), cytokeratin 19 (CK19), human epithelial glycoprotein (EGP) and fibronectin 1 (FN1). Using quadratic discriminant analysis (QDA), expression values were combined into a single score, which indicated CTC-positivity or -negativity. Test characteristics were assessed using receiver operating characteristic (ROC) curve analysis. RESULTS ROC curve analysis showed capability of discrimination between advanced NSCLC patients and healthy controls (area=0.712; 95% CI 0.606-0.819; P<0.001). A cut-off minimizing overall misclassification for QDA-positivity reached a sensitivity of 46% (95% CI 31-61) and a specificity of 93% (95% CI 82-99). CONCLUSIONS In this exploratory study, an assay was developed for discriminating CTCs in peripheral blood samples of advanced NSCLC patients from healthy controls. The assay demonstrated an acceptable sensitivity in combination with good specificity. Further validation studies should take place in NSCLC patients and a matched control group.

Circulating tumor cells as pharmacodynamic biomarker in early clinical oncological trials

Circulating tumor cells (CTCs) have received a lot of attention from both researchers and clinicians because of their prognostic value for progression-free and overall survival in selected tumor types. CTCs are readily available by single venipuncture, thereby posing little burden on the patient and allowing for repeated, sequential sampling during therapy. Nowadays, the sensitivity of several CTC detection and capture techniques allow for further characterization and analysis of specific targets of interest on the CTC itself. These techniques have given CTCs the potential to be used as a pharmacodynamic read-out in drug development. In this review, we explore the utility of CTCs as a pharmacodynamic biomarker in early clinical oncological trials. We present an overview of current literature on assays for CTCs as pharmacodynamic biomarker, their different targets of interest and their level of validation, followed by discussion of their limitations.

Pharmacokinetics of eribulin mesylate in patients with solid tumours receiving repeated oral rifampicin.

Eribulin mesylate is a non‐taxane microtubule dynamics inhibitor that was recently approved for treatment of metastatic breast cancer. The aim of this study was to determine the effect of rifampicin, a CYP3A4 inducer, on the plasma pharmacokinetics of eribulin in patients with solid tumours.

Validation of a multiparameter flow cytometry method for the determination of phosphorylated extracellular-signal-regulated kinase and DNA in circulating tumor cells

A simple, selective, and sensitive multiparameter fluorescence activated cell sorting method utilizing density gradient centrifugation and magnetic antibody cell sorting was developed and validated for the determination of phosphorylated extracellular‐signal‐regulated kinase (pERK) and DNA in circulating tumor cells (CTCs). Cell preparation tubes (CPT) were used for peripheral blood collection and density gradient centrifugation, followed by phosphorylation of ERK with epidermal growth factor (EGF). After fixation with formaldehyde and methanol, magnetic anti epithelial cell adhesion molecule (EpCAM) micro‐beads were used for the selective isolation of CTCs from the background, consisting of peripheral blood mononuclear cells and platelets. Subsequently, samples were stained with Hoechst 33342, and fluorescent antibodies against EpCAM, CD45, and pERK. Flow cytometry was used for identification and enumeration of CTCs and determination of their pERK and DNA content. The validation parameters included specificity, recovery, linearity, precision, sensitivity, and stability. The lower limit of quantification was two CTCs per 8 ml peripheral blood. Samples were stable for 4 months in storage at −80°C. The applicability of the method was demonstrated by successful enumeration of CTCs, and the determination of DNA, and pERK before and after stimulation with EGF in 8 ml peripheral blood samples from patients with metastatic cancer.

Phase I and pharmacological study of pazopanib in combination with oral topotecan in patients with advanced solid tumours.

Background:This phase I study evaluated the safety, tolerability, maximum tolerated dose (MTD) and pharmacokinetics of two dosing schedules of oral topotecan in combination with pazopanib in patients with advanced solid tumours.Methods:Stage I of this study was to determine whether there was an impact of pazopanib on topotecan exposure. In stage II, the MTD and safety profile of oral topotecan given weekly on days 1, 8 and 15 in a 28-day cycle; or daily-times-five on days 1–5 in a 21-day cycle, both in combination with daily pazopanib, were explored.Results:In total, 67 patients were enroled. Pazopanib co-administration caused a substantial increase in exposure to total topotecan (1.7-fold) compared with topotecan alone, which is considered clinically relevant. Topotecan had no effect on pazopanib concentrations. Safety findings were consistent with the known profile of both agents. There were three drug-related deaths, liver failure, tumour haemorrhage and myelosuppression. Two patients experienced dose-limiting toxicities (DLTs; hand–foot syndrome, myelosuppression and diarrhoea) on the weekly topotecan schedule and four patients experienced DLTs (myelosuppression) on the daily-times-five topotecan schedule. When combined with pazopanib, 800 mg daily, the recommended doses for oral topotecan are: 8 mg weekly and 2.5 mg daily-times-five. Seven of eight patients with partial response had platinum-resistant ovarian cancer. In addition, 54% of patients had stable disease with 22% stable for 6 months.Conclusions:Total topotecan exposure is 1.7-fold higher when co-administered with pazopanib. Both schedules of administration were tolerated and would permit further evaluation, especially the weekly schedule.

Pharmacodynamics and pharmacokinetics of oral topotecan in patients with advanced solid tumours and impaired renal function

AIMS The aim of the study was to determine the effect of renal impairment and prior platinum-based chemotherapy on the toxicity and pharmacokinetics of oral topotecan and to identify recommended doses for patients with renal impairment or prior platinum-based (PB) chemotherapy. METHODS A multicentre phase I toxicity and pharmacokinetic study of oral topotecan was conducted in patients with advanced solid tumours. Patients were grouped by normal renal function with limited or prior PB chemotherapy or impaired renal function (mild [creatinine clearance (CLcr)  = 50-79 ml min(-1) ], moderate [CLcr  = 30-49 ml min(-1) ], severe [CLcr <30 ml min(-1) ]). RESULTS Fifty-nine patients were evaluable. Topotecan lactone and total topotecan area under the concentration-time curve (AUC) was significantly increased in patients with moderate and severe renal impairment (109% and 174%, respectively, topotecan lactone and 148% and 298%, respectively, total topotecan). Asian patients (23 in total) had higher AUCs than non-Asian patients with the same degree of renal impairment. Thirteen dose-limiting toxicities (DLTs) were observed, which were mostly haematological. The maximum tolerated dose (MTD) was 2.3 mg m(-2) day(-1) , given on days 1 to 5 in a 21 day cycle, for patients with prior PB chemotherapy or mild renal impairment, and 1.2 mg m(-2) day(-1) for patients with moderate renal impairment (suggested dose 1.9 mg m(-2) day(-1) for non-Asians). Due to incomplete enrolment of patients with severe renal impairment, the MTD was determined as ≥ 0.6 mg m(-2) day(-1) in this cohort. CONCLUSIONS Oral topotecan dose adjustments are not required in patients with prior PB chemotherapy or mildly impaired renal function, but reduced doses are required for patients with moderate or severe renal impairment.