Multi-Marker Immunofluorescent Staining and PD-L1 Detection on Circulating Tumour Cells from Ovarian Cancer Patients
Detection of ovarian cancer (OC) circulating tumour cells (CTCs) is primarily based on targeting epithelial markers, thus failing to detect mesenchymal tumour cells. More importantly, the immune checkpoint inhibitor marker PD-L1 has not been demonstrated on CTCs from OC patients. An antibody stainin...
| Main Authors: | , , , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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MDPI AG
2021-12-01
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| Series: | Cancers |
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| Online Access: | https://www.mdpi.com/2072-6694/13/24/6225 |
| _version_ | 1797506391878926336 |
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| author | Du-Bois Asante Michael Morici Ganendra R. K. A. Mohan Emmanuel Acheampong Isaac Spencer Weitao Lin Paula van Miert Samantha Gibson Aaron B. Beasley Melanie Ziman Leslie Calapre Tarek M. Meniawy Elin S. Gray |
| author_facet | Du-Bois Asante Michael Morici Ganendra R. K. A. Mohan Emmanuel Acheampong Isaac Spencer Weitao Lin Paula van Miert Samantha Gibson Aaron B. Beasley Melanie Ziman Leslie Calapre Tarek M. Meniawy Elin S. Gray |
| author_sort | Du-Bois Asante |
| collection | DOAJ |
| description | Detection of ovarian cancer (OC) circulating tumour cells (CTCs) is primarily based on targeting epithelial markers, thus failing to detect mesenchymal tumour cells. More importantly, the immune checkpoint inhibitor marker PD-L1 has not been demonstrated on CTCs from OC patients. An antibody staining protocol was developed and tested using SKOV-3 and OVCA432 OC cell lines. We targeted epithelial (cytokeratin (CK) and EpCAM), mesenchymal (vimentin), and OC-specific (PAX8) markers for detection of CTCs, and CD45/16 and CD31 were used for the exclusion of white blood and vascular endothelial cells, respectively. PD-L1 was used for CTC characterisation. CTCs were enriched using the Parsortix™ system from 16 OC patients. Results revealed the presence of CTCs in 10 (63%) cases. CTCs were heterogeneous, with 113/157 (72%) cells positive for CK/EpCAM (epithelial marker), 58/157 (37%) positive for vimentin (mesenchymal marker), and 17/157 (11%) for both (hybrid). PAX8 was only found in 11/157 (7%) CTCs. In addition, 62/157 (39%) CTCs were positive for PD-L1. Positivity for PD-L1 was significantly associated with the hybrid phenotype when compared with the epithelial (<i>p</i> = 0.007) and mesenchymal (<i>p</i> = 0.0009) expressing CTCs. Characterisation of CTC phenotypes in relation to clinical outcomes is needed to provide insight into the role that epithelial to mesenchymal plasticity plays in OC and its relationship with PD-L1. |
| first_indexed | 2024-03-10T04:32:01Z |
| format | Article |
| id | doaj.art-a5836bdc61db48d5a75568d57089e7e2 |
| institution | Directory Open Access Journal |
| issn | 2072-6694 |
| language | English |
| last_indexed | 2024-03-10T04:32:01Z |
| publishDate | 2021-12-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Cancers |
| spelling | doaj.art-a5836bdc61db48d5a75568d57089e7e22023-11-23T04:05:31ZengMDPI AGCancers2072-66942021-12-011324622510.3390/cancers13246225Multi-Marker Immunofluorescent Staining and PD-L1 Detection on Circulating Tumour Cells from Ovarian Cancer PatientsDu-Bois Asante0Michael Morici1Ganendra R. K. A. Mohan2Emmanuel Acheampong3Isaac Spencer4Weitao Lin5Paula van Miert6Samantha Gibson7Aaron B. Beasley8Melanie Ziman9Leslie Calapre10Tarek M. Meniawy11Elin S. Gray12School of Medical and Health Sciences, Edith Cowan University, Perth, WA 6027, AustraliaSchool of Medical and Health Sciences, Edith Cowan University, Perth, WA 6027, AustraliaHollywood Private Hospital, Perth, WA 6009, AustraliaSchool of Medical and Health Sciences, Edith Cowan University, Perth, WA 6027, AustraliaSchool of Medical and Health Sciences, Edith Cowan University, Perth, WA 6027, AustraliaSchool of Medical and Health Sciences, Edith Cowan University, Perth, WA 6027, AustraliaSchool of Medical and Health Sciences, Edith Cowan University, Perth, WA 6027, AustraliaWestern Oncology, Perth, WA 6008, AustraliaSchool of Medical and Health Sciences, Edith Cowan University, Perth, WA 6027, AustraliaSchool of Medical and Health Sciences, Edith Cowan University, Perth, WA 6027, AustraliaSchool of Medical and Health Sciences, Edith Cowan University, Perth, WA 6027, AustraliaSchool of Medical and Health Sciences, Edith Cowan University, Perth, WA 6027, AustraliaSchool of Medical and Health Sciences, Edith Cowan University, Perth, WA 6027, AustraliaDetection of ovarian cancer (OC) circulating tumour cells (CTCs) is primarily based on targeting epithelial markers, thus failing to detect mesenchymal tumour cells. More importantly, the immune checkpoint inhibitor marker PD-L1 has not been demonstrated on CTCs from OC patients. An antibody staining protocol was developed and tested using SKOV-3 and OVCA432 OC cell lines. We targeted epithelial (cytokeratin (CK) and EpCAM), mesenchymal (vimentin), and OC-specific (PAX8) markers for detection of CTCs, and CD45/16 and CD31 were used for the exclusion of white blood and vascular endothelial cells, respectively. PD-L1 was used for CTC characterisation. CTCs were enriched using the Parsortix™ system from 16 OC patients. Results revealed the presence of CTCs in 10 (63%) cases. CTCs were heterogeneous, with 113/157 (72%) cells positive for CK/EpCAM (epithelial marker), 58/157 (37%) positive for vimentin (mesenchymal marker), and 17/157 (11%) for both (hybrid). PAX8 was only found in 11/157 (7%) CTCs. In addition, 62/157 (39%) CTCs were positive for PD-L1. Positivity for PD-L1 was significantly associated with the hybrid phenotype when compared with the epithelial (<i>p</i> = 0.007) and mesenchymal (<i>p</i> = 0.0009) expressing CTCs. Characterisation of CTC phenotypes in relation to clinical outcomes is needed to provide insight into the role that epithelial to mesenchymal plasticity plays in OC and its relationship with PD-L1.https://www.mdpi.com/2072-6694/13/24/6225high grade serous ovarian cancerHGSOCimmunofluorescencecirculating tumour cellsCTCfluorescence quenching |
| spellingShingle | Du-Bois Asante Michael Morici Ganendra R. K. A. Mohan Emmanuel Acheampong Isaac Spencer Weitao Lin Paula van Miert Samantha Gibson Aaron B. Beasley Melanie Ziman Leslie Calapre Tarek M. Meniawy Elin S. Gray Multi-Marker Immunofluorescent Staining and PD-L1 Detection on Circulating Tumour Cells from Ovarian Cancer Patients Cancers high grade serous ovarian cancer HGSOC immunofluorescence circulating tumour cells CTC fluorescence quenching |
| title | Multi-Marker Immunofluorescent Staining and PD-L1 Detection on Circulating Tumour Cells from Ovarian Cancer Patients |
| title_full | Multi-Marker Immunofluorescent Staining and PD-L1 Detection on Circulating Tumour Cells from Ovarian Cancer Patients |
| title_fullStr | Multi-Marker Immunofluorescent Staining and PD-L1 Detection on Circulating Tumour Cells from Ovarian Cancer Patients |
| title_full_unstemmed | Multi-Marker Immunofluorescent Staining and PD-L1 Detection on Circulating Tumour Cells from Ovarian Cancer Patients |
| title_short | Multi-Marker Immunofluorescent Staining and PD-L1 Detection on Circulating Tumour Cells from Ovarian Cancer Patients |
| title_sort | multi marker immunofluorescent staining and pd l1 detection on circulating tumour cells from ovarian cancer patients |
| topic | high grade serous ovarian cancer HGSOC immunofluorescence circulating tumour cells CTC fluorescence quenching |
| url | https://www.mdpi.com/2072-6694/13/24/6225 |
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