Digital quantification of somatostatin receptor subtype 2a immunostaining: a validation study

Claudia Campana; Peter M. van Koetsveld; Richard A. Feelders; Wouter W. de Herder; Anand M. Iyer; Marie Louise F. van Velthuysen; Marije J. Veenstra; Elisabeth S.R. van den Dungen; Sanne E. Franck; Diego Ferone; Federico Gatto; Leo J. Hofland

doi:10.1530/EJE-22-0339

Digital quantification of somatostatin receptor subtype 2a immunostaining: a validation study

Claudia Campana, Peter M. van Koetsveld, Richard A. Feelders, Wouter W. de Herder, Anand M. Iyer, Marie Louise F. van Velthuysen, Marije J. Veenstra, Elisabeth S.R. van den Dungen, Sanne E. Franck, Diego Ferone, Federico Gatto, Leo J. Hofland^*

^*Corresponding author for this work

Research output: Contribution to journal › Article › Academic › peer-review

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Abstract

Objective: The aim of this study was to develop an open-source and reproducible digital quantitative analysis (DIA) of somatostatin receptor subtype 2a (SST2) staining in formalin-fixed paraffin-embedded tissues of pancreatic neuroendocrine tumors (panNETs) and growth hormone (GH)-secreting pituitary adenomas (GHomas). Design: SST2 immunostaining of 18 panNETs and 39 GHomas was assessed using a novel DIA protocol and compared with a widely used semi-quantitative immunoreactivity score (IRS). Methods: The DIA software calculates the staining intensity/area and the percentage of positive cells (%PC). Four representative images were selected for each sample by two independent selectors (S1 and S2), with the analysis performed by two independent analyzers (A1 and A2). Agreement between observers was calculated using the concordance correlation coefficient (CCC). Results: In panNETs, the CCC ranged 0.935-0.977 for intensity/area and 0.942-0.983 for %PC. In GHomas, the CCC ranged 0.963-0.997 for intensity/area and 0.979-0.990 for %PC. In both panNETs and GHomas, the DIA staining intensity was strongly correlated with the IRS (Spearman rho: 0.916-0.969, P < 0.001), as well as the DIA %PC with the IRS %PC (Spearman rh: 0.826-0.881, P < 0.001). In GHomas, the biochemical response to somatostatin receptor ligands correlated with SST2 expression, evaluated both as DIA intensity/area (Spearman rho: -0.448 to -0.527, P = 0.007-0.004) and DIA %PC (Spearman rho: -0.558 to -0.644, P ≤ 0.001). Conclusions: The DIA has an excellent inter-observer agreement and showed a strong correlation with the widely used semi-quantitative IRS. The DIA protocol is an open-source, highly reproducible tool and provides a reliable quantitative evaluation of SST2 immunohistochemistry.

Original language	English
Pages (from-to)	399-411
Number of pages	13
Journal	European Journal of Endocrinology
Volume	187
Issue number	3
DOIs	https://doi.org/10.1530/EJE-22-0339
Publication status	Published - 21 Jul 2022

Bibliographical note

Funding Information:
R A F has received research grants from Strongbridge and Corcept and is a consultant for Recordati. W W H has been a speaker for and participated on advisory boards and received research grants from Novartis and Ipsen. D F has been a speaker for and participated on advisory boards and received research grants from Novartis-AAA, Ipsen, Recordati RD, Camurus and Pfizer. F G has been a speaker for Novartis and has participated on advisory boards of Novartis, AMCo Ltd, and IONIS Pharmaceuticals. The other authors have no conflict of interest to declare.

Publisher Copyright:
© 2022 The authors.

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[1479683X - European Journal of Endocrinology] Digital quantification of somatostatin receptor subtype 2a immunostaining_ a validation studyFinal published version, 2.32 MBLicence: CC BY

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Campana, C., van Koetsveld, P. M., Feelders, R. A., de Herder, W. W., Iyer, A. M., van Velthuysen, M. L. F., Veenstra, M. J., van den Dungen, E. S. R., Franck, S. E., Ferone, D., Gatto, F., & Hofland, L. J. (2022). Digital quantification of somatostatin receptor subtype 2a immunostaining: a validation study. European Journal of Endocrinology, 187(3), 399-411. https://doi.org/10.1530/EJE-22-0339

@article{3353ad0e416a4d00acba7f9623cf6511,

title = "Digital quantification of somatostatin receptor subtype 2a immunostaining: a validation study",

abstract = "Objective: The aim of this study was to develop an open-source and reproducible digital quantitative analysis (DIA) of somatostatin receptor subtype 2a (SST2) staining in formalin-fixed paraffin-embedded tissues of pancreatic neuroendocrine tumors (panNETs) and growth hormone (GH)-secreting pituitary adenomas (GHomas). Design: SST2 immunostaining of 18 panNETs and 39 GHomas was assessed using a novel DIA protocol and compared with a widely used semi-quantitative immunoreactivity score (IRS). Methods: The DIA software calculates the staining intensity/area and the percentage of positive cells (%PC). Four representative images were selected for each sample by two independent selectors (S1 and S2), with the analysis performed by two independent analyzers (A1 and A2). Agreement between observers was calculated using the concordance correlation coefficient (CCC). Results: In panNETs, the CCC ranged 0.935-0.977 for intensity/area and 0.942-0.983 for %PC. In GHomas, the CCC ranged 0.963-0.997 for intensity/area and 0.979-0.990 for %PC. In both panNETs and GHomas, the DIA staining intensity was strongly correlated with the IRS (Spearman rho: 0.916-0.969, P < 0.001), as well as the DIA %PC with the IRS %PC (Spearman rh: 0.826-0.881, P < 0.001). In GHomas, the biochemical response to somatostatin receptor ligands correlated with SST2 expression, evaluated both as DIA intensity/area (Spearman rho: -0.448 to -0.527, P = 0.007-0.004) and DIA %PC (Spearman rho: -0.558 to -0.644, P ≤ 0.001). Conclusions: The DIA has an excellent inter-observer agreement and showed a strong correlation with the widely used semi-quantitative IRS. The DIA protocol is an open-source, highly reproducible tool and provides a reliable quantitative evaluation of SST2 immunohistochemistry.",

author = "Claudia Campana and {van Koetsveld}, {Peter M.} and Feelders, {Richard A.} and {de Herder}, {Wouter W.} and Iyer, {Anand M.} and {van Velthuysen}, {Marie Louise F.} and Veenstra, {Marije J.} and {van den Dungen}, {Elisabeth S.R.} and Franck, {Sanne E.} and Diego Ferone and Federico Gatto and Hofland, {Leo J.}",

note = "Funding Information: R A F has received research grants from Strongbridge and Corcept and is a consultant for Recordati. W W H has been a speaker for and participated on advisory boards and received research grants from Novartis and Ipsen. D F has been a speaker for and participated on advisory boards and received research grants from Novartis-AAA, Ipsen, Recordati RD, Camurus and Pfizer. F G has been a speaker for Novartis and has participated on advisory boards of Novartis, AMCo Ltd, and IONIS Pharmaceuticals. The other authors have no conflict of interest to declare. Publisher Copyright: {\textcopyright} 2022 The authors.",

year = "2022",

month = jul,

day = "21",

doi = "10.1530/EJE-22-0339",

language = "English",

volume = "187",

pages = "399--411",

journal = "European Journal of Endocrinology",

issn = "0804-4643",

publisher = "Bioscientifica Ltd",

number = "3",

}

Campana, C , van Koetsveld, PM , Feelders, RA , de Herder, WW , Iyer, AM , van Velthuysen, MLF, Veenstra, MJ, van den Dungen, ESR, Franck, SE, Ferone, D, Gatto, F & Hofland, LJ 2022, 'Digital quantification of somatostatin receptor subtype 2a immunostaining: a validation study', European Journal of Endocrinology, vol. 187, no. 3, pp. 399-411. https://doi.org/10.1530/EJE-22-0339

TY - JOUR

T1 - Digital quantification of somatostatin receptor subtype 2a immunostaining

T2 - a validation study

AU - Campana, Claudia

AU - van Koetsveld, Peter M.

AU - Feelders, Richard A.

AU - de Herder, Wouter W.

AU - Iyer, Anand M.

AU - van Velthuysen, Marie Louise F.

AU - Veenstra, Marije J.

AU - van den Dungen, Elisabeth S.R.

AU - Franck, Sanne E.

AU - Ferone, Diego

AU - Gatto, Federico

AU - Hofland, Leo J.

N1 - Funding Information: R A F has received research grants from Strongbridge and Corcept and is a consultant for Recordati. W W H has been a speaker for and participated on advisory boards and received research grants from Novartis and Ipsen. D F has been a speaker for and participated on advisory boards and received research grants from Novartis-AAA, Ipsen, Recordati RD, Camurus and Pfizer. F G has been a speaker for Novartis and has participated on advisory boards of Novartis, AMCo Ltd, and IONIS Pharmaceuticals. The other authors have no conflict of interest to declare. Publisher Copyright: © 2022 The authors.

PY - 2022/7/21

Y1 - 2022/7/21

N2 - Objective: The aim of this study was to develop an open-source and reproducible digital quantitative analysis (DIA) of somatostatin receptor subtype 2a (SST2) staining in formalin-fixed paraffin-embedded tissues of pancreatic neuroendocrine tumors (panNETs) and growth hormone (GH)-secreting pituitary adenomas (GHomas). Design: SST2 immunostaining of 18 panNETs and 39 GHomas was assessed using a novel DIA protocol and compared with a widely used semi-quantitative immunoreactivity score (IRS). Methods: The DIA software calculates the staining intensity/area and the percentage of positive cells (%PC). Four representative images were selected for each sample by two independent selectors (S1 and S2), with the analysis performed by two independent analyzers (A1 and A2). Agreement between observers was calculated using the concordance correlation coefficient (CCC). Results: In panNETs, the CCC ranged 0.935-0.977 for intensity/area and 0.942-0.983 for %PC. In GHomas, the CCC ranged 0.963-0.997 for intensity/area and 0.979-0.990 for %PC. In both panNETs and GHomas, the DIA staining intensity was strongly correlated with the IRS (Spearman rho: 0.916-0.969, P < 0.001), as well as the DIA %PC with the IRS %PC (Spearman rh: 0.826-0.881, P < 0.001). In GHomas, the biochemical response to somatostatin receptor ligands correlated with SST2 expression, evaluated both as DIA intensity/area (Spearman rho: -0.448 to -0.527, P = 0.007-0.004) and DIA %PC (Spearman rho: -0.558 to -0.644, P ≤ 0.001). Conclusions: The DIA has an excellent inter-observer agreement and showed a strong correlation with the widely used semi-quantitative IRS. The DIA protocol is an open-source, highly reproducible tool and provides a reliable quantitative evaluation of SST2 immunohistochemistry.

AB - Objective: The aim of this study was to develop an open-source and reproducible digital quantitative analysis (DIA) of somatostatin receptor subtype 2a (SST2) staining in formalin-fixed paraffin-embedded tissues of pancreatic neuroendocrine tumors (panNETs) and growth hormone (GH)-secreting pituitary adenomas (GHomas). Design: SST2 immunostaining of 18 panNETs and 39 GHomas was assessed using a novel DIA protocol and compared with a widely used semi-quantitative immunoreactivity score (IRS). Methods: The DIA software calculates the staining intensity/area and the percentage of positive cells (%PC). Four representative images were selected for each sample by two independent selectors (S1 and S2), with the analysis performed by two independent analyzers (A1 and A2). Agreement between observers was calculated using the concordance correlation coefficient (CCC). Results: In panNETs, the CCC ranged 0.935-0.977 for intensity/area and 0.942-0.983 for %PC. In GHomas, the CCC ranged 0.963-0.997 for intensity/area and 0.979-0.990 for %PC. In both panNETs and GHomas, the DIA staining intensity was strongly correlated with the IRS (Spearman rho: 0.916-0.969, P < 0.001), as well as the DIA %PC with the IRS %PC (Spearman rh: 0.826-0.881, P < 0.001). In GHomas, the biochemical response to somatostatin receptor ligands correlated with SST2 expression, evaluated both as DIA intensity/area (Spearman rho: -0.448 to -0.527, P = 0.007-0.004) and DIA %PC (Spearman rho: -0.558 to -0.644, P ≤ 0.001). Conclusions: The DIA has an excellent inter-observer agreement and showed a strong correlation with the widely used semi-quantitative IRS. The DIA protocol is an open-source, highly reproducible tool and provides a reliable quantitative evaluation of SST2 immunohistochemistry.

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U2 - 10.1530/EJE-22-0339

DO - 10.1530/EJE-22-0339

M3 - Article

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AN - SCOPUS:85135374690

SN - 0804-4643

VL - 187

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EP - 411

JO - European Journal of Endocrinology

JF - European Journal of Endocrinology

IS - 3

ER -

Digital quantification of somatostatin receptor subtype 2a immunostaining: a validation study

Abstract

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