Direct Chemosensitivity Monitoring Ex Vivo on Undissociated Melanoma Tumor Tissue by Impedance Spectroscopy

Abstract

Stage III/IV melanoma remains incurable in most cases due to chemotherapeutic resistance. Thus, predicting and monitoring chemotherapeutic responses in this setting offer great interest. To overcome limitations of existing assays in evaluating the chemosensitivity of dissociated tumor cells, we developed a label-free monitoring system to directly analyze the chemosensitivity of undissociated tumor tissue. Using a preparation of tumor micro-fragments (TMF) established from melanoma biopsies, we characterized the tissue organization and biomarker expression by immunocytochemistry. Robust generation of TMF was established successfully and demonstrated on a broad range of primary melanoma tumors and tumor metastases. Organization and biomarker expression within the TMF were highly comparable with tumor tissue, in contrast to dissociated, cultivated tumor cells. Using isolated TMF, sensitivity to six clinically relevant chemotherapeutic drugs (dacarbazine, doxorubicin, paclitaxel, cisplatin, gemcitabine, and treosulfan) was determined by impedance spectroscopy in combination with a unique microcavity array technology we developed. In parallel, comparative analyses were performed on monolayer tumor cell cultures. Lastly, we determined the efficacy of chemotherapeutic agents on TMF by impedance spectroscopy to obtain individual chemosensitivity patterns. Our results demonstrated nonpredictable differences in the reaction of tumor cells to chemotherapy in TMF by comparison with dissociated, cultivated tumor cells. Our direct impedimetric analysis of melanoma biopsies offers a direct ex vivo system to more reliably predict patient-specific chemosensitivity patterns and to monitor antitumor efficacy. Cancer Res; 74(22); 6408-18. Ó2014 AACR.

Induced cross-resistance of BRAFV600E melanoma cells to standard chemotherapeutic dacarbazine after chronic PLX4032 treatment

“…While commonly applied XTT or MTT in vitro cytotoxicity assays have a limited ability to detect cytostatic effects 46 , our non-invasive, label-free, real-time impedance technology allows precise and comprehensive chemosensitivity analysis in terms of both, cytotoxic and cytostatic compound screening 18–20 . Using impedance spectroscopy, we could identify time- and concentration-dependent cross-desensitisation patterns towards chemotherapeutic dacarbazine and BRAF V600E -targeting drug PLX4032 in BRAF-mutated metastatic melanoma cells that clearly correlated with the underlying activation/inhibition of MAPK- and PI3K/AKT survival signalling.…”

“…Until now, no study exists that examines the interrelation in resistance acquisition of currently applied oncogene targeting therapeutics and adjacent classical chemotherapy, which severely interferes with patients’ long-term survival. Here, we focus on the analysis of cross-resistance patterns in BRAF-mutated melanoma cells using microelectrode array-based impedance spectroscopy, a non-invasive, label-free bioelectronic method that was recently validated over standard XTT and ATP assays for the sensitive and comprehensive real-time detection of cellular drug effects in vitro 18–20 .…”

Precision medicine for cancer with next-generation functional diagnostics

“…Correlation with patient outcomes will be needed to confirm that these overactive targets and pathways in patient biopsies are real vulnerabilities or ‘drivers’. Many other novel assays that provide functional readouts from live patient biopsies have been described in the past few years, including impedance spectroscopy to measure ex vivo responses of fragments of live melanoma tumour specimens to chemotherapy 82 , and metabolic outputs such as oxygen consumption 83 . Similarly to measurements of proliferation, viability and apoptosis, these assays will require rigorous evaluation of their relevance to prediction of patient outcomes and comparison between them to determine the advantages and disadvantages of each assay beyond technical differences in reagents, timing and analysis.…”

Electrochemical live monitoring of tumor cell migration out of micro-tumors on an innovative multiwell high-dense microelectrode array

“…As a reference for a highly migrating tumor cell line, MDA-MB-231 cells were chosen 26,27 . Additionally, we investigated two self-established melanoma cell lines (T12.8.10ZII and T30.6.9) 28 that seem to have at least slightly different migration characteristics from microscopic observations. Initially, our micro-tumor models were characterized concerning spheroid size as well as impedimetric analysis of spheroid density with our self-developed microcavity array 28,29 .…”