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Real-time Detection of Breast Cancer Cells Using Peptide-functionalized Microcantilever Arrays.

Etayash H, Jiang K, Azmi S, Thundat T, Kaur K - Sci Rep (2015)

Bottom Line: Ligands such as antibodies have been successfully used for capturing cancer cells and an antibody based system (CellSearch(®)) is currently used clinically to enumerate CTCs.A sensing microcantilever, functionalized with a breast cancer specific peptide 18-4 (WxEAAYQrFL), showed significant deflection on cancer cell (MCF7 and MDA-MB-231) binding compared to when exposed to noncancerous (MCF10A and HUVEC) cells.The reported peptide-based cantilever platform represents a new analytical approach that can lead to an alternative to the various detection platforms and can be leveraged to further study CTCs.

View Article: PubMed Central - PubMed

Affiliation: Faculty of Pharmacy and Pharmaceutical Sciences, University of Alberta, Edmonton, Alberta, T6G 2E1, Canada.

ABSTRACT
Ligand-directed targeting and capturing of cancer cells is a new approach for detecting circulating tumor cells (CTCs). Ligands such as antibodies have been successfully used for capturing cancer cells and an antibody based system (CellSearch(®)) is currently used clinically to enumerate CTCs. Here we report the use of a peptide moiety in conjunction with a microcantilever array system to selectively detect CTCs resulting from cancer, specifically breast cancer. A sensing microcantilever, functionalized with a breast cancer specific peptide 18-4 (WxEAAYQrFL), showed significant deflection on cancer cell (MCF7 and MDA-MB-231) binding compared to when exposed to noncancerous (MCF10A and HUVEC) cells. The peptide-functionalized microcantilever allowed efficient capture and detection of cancer cells in MCF7 spiked human blood samples emulating CTCs in human blood. A detection limit of 50-100 cancer cells mL(-1) from blood samples was achieved with a capture yield of 80% from spiked whole blood samples. The results emphasize the potential of peptide 18-4 as a novel peptide for capturing and detecting cancer cells in conjunction with nanomechanical cantilever platform. The reported peptide-based cantilever platform represents a new analytical approach that can lead to an alternative to the various detection platforms and can be leveraged to further study CTCs.

No MeSH data available.


Related in: MedlinePlus

Specific binding of peptide-cantilevers to breast cancer cells.(a) Label-free real time recognition of cancer cell lines (MCF7 or MDA-MB-231, 100 ± 10 cells mL−1) from non-cancerous cell lines (MCF10A or HUVEC, 100 ± 10 cells mL−1) with microcantilever array functionalized with 18-4 cancer targeting peptide. (b) Cantilever deflection in response to a function of different concentration ratios of cancerous to non-cancerous cells (MCF7 to MCF10A) in PBS solution as indicated. (c) and (d) demonstrate the concentration dependence of cantilever response to the number of cancerous or non-cancerous cells, respectively, present in the co-culture sample (100 cells mL−1). The representative graphs show an increase in cantilever deflection with an increase in the number of MCF7 cells (c), and a decrease in cantilever deflection with an increase in the number of MCF10A (d) in the media. Each cantilever differential deflection represents an average calculation of eight replicates and error bars indicate standard deviations.
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f3: Specific binding of peptide-cantilevers to breast cancer cells.(a) Label-free real time recognition of cancer cell lines (MCF7 or MDA-MB-231, 100 ± 10 cells mL−1) from non-cancerous cell lines (MCF10A or HUVEC, 100 ± 10 cells mL−1) with microcantilever array functionalized with 18-4 cancer targeting peptide. (b) Cantilever deflection in response to a function of different concentration ratios of cancerous to non-cancerous cells (MCF7 to MCF10A) in PBS solution as indicated. (c) and (d) demonstrate the concentration dependence of cantilever response to the number of cancerous or non-cancerous cells, respectively, present in the co-culture sample (100 cells mL−1). The representative graphs show an increase in cantilever deflection with an increase in the number of MCF7 cells (c), and a decrease in cantilever deflection with an increase in the number of MCF10A (d) in the media. Each cantilever differential deflection represents an average calculation of eight replicates and error bars indicate standard deviations.

Mentions: In order to determine the specificity of the designed sensor, we applied the peptide 18-4 functionalized cantilever array to distinguish between cancerous and non-cancerous cell lines in real-time (Fig. 3). Here the binding affinity of the peptide sensor was explored against two types of breast cancer cell lines, namely, MCF7 and MDA-MB-231 and two non-cancerous cell lines, MCF10A and HUVEC. MCF10A are non-cancerous cells derived from the same human mammary tissue as MCF7, whereas HUVEC are endothelial cells isolated from normal human umbilical vein. When cells were injected at a concentration of 100 ± 10 cells mL−1 separately to each peptide cantilever, the cantilever showed significant deflection for cancerous cell binding (280 ± 25 nm) compared to non-cancerous cell binding (90 ± 15 nm, Fig. 3a). The variation in cantilever deflection upon binding cancerous or noncancerous cells is most likely due to the differential expression levels of specific peptide-binding receptors present in cancerous and noncancerous cells. We and others have shown that peptide 18-4 and the original lead peptide p160 enter cells by a receptor-mediated endocytosis2123. The receptor is not known yet, however, it is clear that the receptor is overexpressed in breast cancer cells compared to normal cells. The results confirm our conjecture that peptide 18-4 binds breast cancer cells with high specificity. Previously we showed that a similar peptide, peptide 18 (WXEAAYQRFL), binds MDA-MB-435 breast cancer cells with an apparent Kd of 41.9 μM22.


Real-time Detection of Breast Cancer Cells Using Peptide-functionalized Microcantilever Arrays.

Etayash H, Jiang K, Azmi S, Thundat T, Kaur K - Sci Rep (2015)

Specific binding of peptide-cantilevers to breast cancer cells.(a) Label-free real time recognition of cancer cell lines (MCF7 or MDA-MB-231, 100 ± 10 cells mL−1) from non-cancerous cell lines (MCF10A or HUVEC, 100 ± 10 cells mL−1) with microcantilever array functionalized with 18-4 cancer targeting peptide. (b) Cantilever deflection in response to a function of different concentration ratios of cancerous to non-cancerous cells (MCF7 to MCF10A) in PBS solution as indicated. (c) and (d) demonstrate the concentration dependence of cantilever response to the number of cancerous or non-cancerous cells, respectively, present in the co-culture sample (100 cells mL−1). The representative graphs show an increase in cantilever deflection with an increase in the number of MCF7 cells (c), and a decrease in cantilever deflection with an increase in the number of MCF10A (d) in the media. Each cantilever differential deflection represents an average calculation of eight replicates and error bars indicate standard deviations.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4593050&req=5

f3: Specific binding of peptide-cantilevers to breast cancer cells.(a) Label-free real time recognition of cancer cell lines (MCF7 or MDA-MB-231, 100 ± 10 cells mL−1) from non-cancerous cell lines (MCF10A or HUVEC, 100 ± 10 cells mL−1) with microcantilever array functionalized with 18-4 cancer targeting peptide. (b) Cantilever deflection in response to a function of different concentration ratios of cancerous to non-cancerous cells (MCF7 to MCF10A) in PBS solution as indicated. (c) and (d) demonstrate the concentration dependence of cantilever response to the number of cancerous or non-cancerous cells, respectively, present in the co-culture sample (100 cells mL−1). The representative graphs show an increase in cantilever deflection with an increase in the number of MCF7 cells (c), and a decrease in cantilever deflection with an increase in the number of MCF10A (d) in the media. Each cantilever differential deflection represents an average calculation of eight replicates and error bars indicate standard deviations.
Mentions: In order to determine the specificity of the designed sensor, we applied the peptide 18-4 functionalized cantilever array to distinguish between cancerous and non-cancerous cell lines in real-time (Fig. 3). Here the binding affinity of the peptide sensor was explored against two types of breast cancer cell lines, namely, MCF7 and MDA-MB-231 and two non-cancerous cell lines, MCF10A and HUVEC. MCF10A are non-cancerous cells derived from the same human mammary tissue as MCF7, whereas HUVEC are endothelial cells isolated from normal human umbilical vein. When cells were injected at a concentration of 100 ± 10 cells mL−1 separately to each peptide cantilever, the cantilever showed significant deflection for cancerous cell binding (280 ± 25 nm) compared to non-cancerous cell binding (90 ± 15 nm, Fig. 3a). The variation in cantilever deflection upon binding cancerous or noncancerous cells is most likely due to the differential expression levels of specific peptide-binding receptors present in cancerous and noncancerous cells. We and others have shown that peptide 18-4 and the original lead peptide p160 enter cells by a receptor-mediated endocytosis2123. The receptor is not known yet, however, it is clear that the receptor is overexpressed in breast cancer cells compared to normal cells. The results confirm our conjecture that peptide 18-4 binds breast cancer cells with high specificity. Previously we showed that a similar peptide, peptide 18 (WXEAAYQRFL), binds MDA-MB-435 breast cancer cells with an apparent Kd of 41.9 μM22.

Bottom Line: Ligands such as antibodies have been successfully used for capturing cancer cells and an antibody based system (CellSearch(®)) is currently used clinically to enumerate CTCs.A sensing microcantilever, functionalized with a breast cancer specific peptide 18-4 (WxEAAYQrFL), showed significant deflection on cancer cell (MCF7 and MDA-MB-231) binding compared to when exposed to noncancerous (MCF10A and HUVEC) cells.The reported peptide-based cantilever platform represents a new analytical approach that can lead to an alternative to the various detection platforms and can be leveraged to further study CTCs.

View Article: PubMed Central - PubMed

Affiliation: Faculty of Pharmacy and Pharmaceutical Sciences, University of Alberta, Edmonton, Alberta, T6G 2E1, Canada.

ABSTRACT
Ligand-directed targeting and capturing of cancer cells is a new approach for detecting circulating tumor cells (CTCs). Ligands such as antibodies have been successfully used for capturing cancer cells and an antibody based system (CellSearch(®)) is currently used clinically to enumerate CTCs. Here we report the use of a peptide moiety in conjunction with a microcantilever array system to selectively detect CTCs resulting from cancer, specifically breast cancer. A sensing microcantilever, functionalized with a breast cancer specific peptide 18-4 (WxEAAYQrFL), showed significant deflection on cancer cell (MCF7 and MDA-MB-231) binding compared to when exposed to noncancerous (MCF10A and HUVEC) cells. The peptide-functionalized microcantilever allowed efficient capture and detection of cancer cells in MCF7 spiked human blood samples emulating CTCs in human blood. A detection limit of 50-100 cancer cells mL(-1) from blood samples was achieved with a capture yield of 80% from spiked whole blood samples. The results emphasize the potential of peptide 18-4 as a novel peptide for capturing and detecting cancer cells in conjunction with nanomechanical cantilever platform. The reported peptide-based cantilever platform represents a new analytical approach that can lead to an alternative to the various detection platforms and can be leveraged to further study CTCs.

No MeSH data available.


Related in: MedlinePlus