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BMC Cancer
Published in: BMC Cancer 1/2015

Open Access 01-12-2015 | Technical advance

Optimisation of immunofluorescence methods to determine MCT1 and MCT4 expression in circulating tumour cells

Authors: Stephen Kershaw, Jeffrey Cummings, Karen Morris, Jonathan Tugwood, Caroline Dive

Published in: BMC Cancer | Issue 1/2015

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Abstract

Background

The monocarboxylate transporter-1 (MCT1) represents a novel target in rational anticancer drug design while AZD3965 was developed as an inhibitor of this transporter and is undergoing Phase I clinical trials (http://​www.​clinicaltrials.​gov/​show/​NCT01791595). We describe the optimisation of an immunofluorescence (IF) method for determination of MCT1 and MCT4 in circulating tumour cells (CTC) as potential prognostic and predictive biomarkers of AZD3965 in cancer patients.

Methods

Antibody selectivity was investigated by western blotting (WB) in K562 and MDAMB231 cell lines acting as positive controls for MCT1 and MCT4 respectively and by flow cytometry also employing the control cell lines. Ability to detect MCT1 and MCT4 in CTC as a 4th channel marker utilising the Veridex™ CellSearch system was conducted in both human volunteer blood spiked with control cells and in samples collected from small cell lung cancer (SCLC) patients.

Results

Experimental conditions were established which yielded a 10-fold dynamic range (DR) for detection of MCT1 over MCT4 (antibody concentration 6.25 μg/mL; integration time 0.4 seconds) and a 5-fold DR of MCT4 over MCT 1 (8 μg/100 μL and 0.8 seconds). The IF method was sufficiently sensitive to detect both MCT1 and MCT4 in CTCs harvested from cancer patients.

Conclusions

The first IF method has been developed and optimised for detection of MCT 1 and MCT4 in cancer patient CTC.
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Metadata
Title
Optimisation of immunofluorescence methods to determine MCT1 and MCT4 expression in circulating tumour cells
Authors
Stephen Kershaw
Jeffrey Cummings
Karen Morris
Jonathan Tugwood
Caroline Dive
Publication date
01-12-2015
Publisher
BioMed Central
Published in
BMC Cancer / Issue 1/2015
Electronic ISSN: 1471-2407
DOI
https://doi.org/10.1186/s12885-015-1382-y

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