Calreticulin mutation specific CAL2 immunohistochemistry accurately identifies rare calreticulin mutations in myeloproliferative neoplasms

Réka Mózes; Ambrus Gángó; Adrienn Sulák; Livia Vida; Lilla Reiniger; Botond Timár; T. Krenács; Hussain Alizadeh; T. Masszi; Júlia Gaál-Weisinger; J. Demeter; Judit Csomor; A. Matolcsy; Béla Kajtár; Csaba Bödör

doi:10.1016/j.pathol.2018.11.007

Calreticulin mutation specific CAL2 immunohistochemistry accurately identifies rare calreticulin mutations in myeloproliferative neoplasms

Réka Mózes, Ambrus Gángó, Adrienn Sulák, Livia Vida, Lilla Reiniger, Botond Timár, T. Krenács, Hussain Alizadeh, T. Masszi, Júlia Gaál-Weisinger, J. Demeter, Judit Csomor, A. Matolcsy, Béla Kajtár, Csaba Bödör

Research output: Contribution to journal › Article

Abstract

Mutations of the multifunctional protein calreticulin (CALR) are recognised as one of the main driver alterations involved in the pathogenesis of Philadelphia negative myeloproliferative neoplasms (Ph^– MPN) and also represent a major diagnostic criterion in the most recent World Health Organization classification of myeloid neoplasms. Nowadays, quantitative assessment of the driver mutations is gaining importance, as recent studies demonstrated the clinical relevance of the mutation load reflecting the size of the mutant clone. Here, we performed for the first time a manual and automated quantitative assessment of the CALR mutation load at protein level using CAL2, a recently developed CALR mutation specific monoclonal antibody, on a cohort of 117 patients with essential thrombocythemia (ET) or primary myelofibrosis (PMF) and compared the CALR protein mutation loads with the CALR mutation load values established by a molecular assay. Eighteen different CALR mutations were detected in the cohort of the 91 CALR mutant cases. Mutation loads of the CALR mutations were between 13% and 94% with mean value in PMF cases significantly higher than ET cases (49.94 vs 41.09; t-test, p=0.004). Cases without CALR mutation (n=26) showed no or only minimal labelling with the CAL2 antibody, while all 18 different types of CALR mutations were associated with CAL2 labelling. The CALR mutation load showed a significant correlation (p=0.03) with the occurrence of major thrombotic events, with higher mutation load in patients presenting with these complications. We report a 100% concordance between the mutation status determined by immunohistochemistry and the CALR molecular assay, and we extend the applicability of this approach to 16 rare CALR mutations previously not analysed at protein level.

Original language	English
Journal	Pathology
DOIs	https://doi.org/10.1016/j.pathol.2018.11.007
Publication status	Accepted/In press - Jan 1 2019

Fingerprint

Calreticulin

Immunohistochemistry

Mutation

Neoplasms

Essential Thrombocythemia

Primary Myelofibrosis

Proteins

Keywords

CAL2 immunohistochemistry
calreticulin mutation
mutation load
Myeloproliferative neoplasms

ASJC Scopus subject areas

Pathology and Forensic Medicine

Cite this

Mózes, R., Gángó, A., Sulák, A., Vida, L., Reiniger, L., Timár, B., ... Bödör, C. (Accepted/In press). Calreticulin mutation specific CAL2 immunohistochemistry accurately identifies rare calreticulin mutations in myeloproliferative neoplasms. Pathology. https://doi.org/10.1016/j.pathol.2018.11.007

Calreticulin mutation specific CAL2 immunohistochemistry accurately identifies rare calreticulin mutations in myeloproliferative neoplasms. / Mózes, Réka; Gángó, Ambrus; Sulák, Adrienn; Vida, Livia; Reiniger, Lilla; Timár, Botond; Krenács, T.; Alizadeh, Hussain; Masszi, T.; Gaál-Weisinger, Júlia; Demeter, J.; Csomor, Judit; Matolcsy, A.; Kajtár, Béla; Bödör, Csaba.

In: Pathology, 01.01.2019.

Research output: Contribution to journal › Article

Mózes, R, Gángó, A, Sulák, A, Vida, L, Reiniger, L, Timár, B, Krenács, T, Alizadeh, H, Masszi, T, Gaál-Weisinger, J, Demeter, J, Csomor, J, Matolcsy, A, Kajtár, B & Bödör, C 2019, 'Calreticulin mutation specific CAL2 immunohistochemistry accurately identifies rare calreticulin mutations in myeloproliferative neoplasms', Pathology. https://doi.org/10.1016/j.pathol.2018.11.007

Mózes R, Gángó A, Sulák A, Vida L, Reiniger L, Timár B et al. Calreticulin mutation specific CAL2 immunohistochemistry accurately identifies rare calreticulin mutations in myeloproliferative neoplasms. Pathology. 2019 Jan 1. https://doi.org/10.1016/j.pathol.2018.11.007

Mózes, Réka ; Gángó, Ambrus ; Sulák, Adrienn ; Vida, Livia ; Reiniger, Lilla ; Timár, Botond ; Krenács, T. ; Alizadeh, Hussain ; Masszi, T. ; Gaál-Weisinger, Júlia ; Demeter, J. ; Csomor, Judit ; Matolcsy, A. ; Kajtár, Béla ; Bödör, Csaba. / Calreticulin mutation specific CAL2 immunohistochemistry accurately identifies rare calreticulin mutations in myeloproliferative neoplasms. In: Pathology. 2019.

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abstract = "Mutations of the multifunctional protein calreticulin (CALR) are recognised as one of the main driver alterations involved in the pathogenesis of Philadelphia negative myeloproliferative neoplasms (Ph– MPN) and also represent a major diagnostic criterion in the most recent World Health Organization classification of myeloid neoplasms. Nowadays, quantitative assessment of the driver mutations is gaining importance, as recent studies demonstrated the clinical relevance of the mutation load reflecting the size of the mutant clone. Here, we performed for the first time a manual and automated quantitative assessment of the CALR mutation load at protein level using CAL2, a recently developed CALR mutation specific monoclonal antibody, on a cohort of 117 patients with essential thrombocythemia (ET) or primary myelofibrosis (PMF) and compared the CALR protein mutation loads with the CALR mutation load values established by a molecular assay. Eighteen different CALR mutations were detected in the cohort of the 91 CALR mutant cases. Mutation loads of the CALR mutations were between 13{\%} and 94{\%} with mean value in PMF cases significantly higher than ET cases (49.94 vs 41.09; t-test, p=0.004). Cases without CALR mutation (n=26) showed no or only minimal labelling with the CAL2 antibody, while all 18 different types of CALR mutations were associated with CAL2 labelling. The CALR mutation load showed a significant correlation (p=0.03) with the occurrence of major thrombotic events, with higher mutation load in patients presenting with these complications. We report a 100{\%} concordance between the mutation status determined by immunohistochemistry and the CALR molecular assay, and we extend the applicability of this approach to 16 rare CALR mutations previously not analysed at protein level.",

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AU - Reiniger, Lilla

AU - Timár, Botond

AU - Krenács, T.

AU - Alizadeh, Hussain

AU - Masszi, T.

AU - Gaál-Weisinger, Júlia

AU - Demeter, J.

AU - Csomor, Judit

AU - Matolcsy, A.

AU - Kajtár, Béla

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AB - Mutations of the multifunctional protein calreticulin (CALR) are recognised as one of the main driver alterations involved in the pathogenesis of Philadelphia negative myeloproliferative neoplasms (Ph– MPN) and also represent a major diagnostic criterion in the most recent World Health Organization classification of myeloid neoplasms. Nowadays, quantitative assessment of the driver mutations is gaining importance, as recent studies demonstrated the clinical relevance of the mutation load reflecting the size of the mutant clone. Here, we performed for the first time a manual and automated quantitative assessment of the CALR mutation load at protein level using CAL2, a recently developed CALR mutation specific monoclonal antibody, on a cohort of 117 patients with essential thrombocythemia (ET) or primary myelofibrosis (PMF) and compared the CALR protein mutation loads with the CALR mutation load values established by a molecular assay. Eighteen different CALR mutations were detected in the cohort of the 91 CALR mutant cases. Mutation loads of the CALR mutations were between 13% and 94% with mean value in PMF cases significantly higher than ET cases (49.94 vs 41.09; t-test, p=0.004). Cases without CALR mutation (n=26) showed no or only minimal labelling with the CAL2 antibody, while all 18 different types of CALR mutations were associated with CAL2 labelling. The CALR mutation load showed a significant correlation (p=0.03) with the occurrence of major thrombotic events, with higher mutation load in patients presenting with these complications. We report a 100% concordance between the mutation status determined by immunohistochemistry and the CALR molecular assay, and we extend the applicability of this approach to 16 rare CALR mutations previously not analysed at protein level.

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10.1016/j.pathol.2018.11.007