Secreted mutant calreticulins as rogue cytokines in myeloproliferative neoplasms

• Published in: Blood Vol. 141; no. 8; pp. 917 - 929
• Main Authors: Pecquet, Christian, Papadopoulos, Nicolas, Balligand, Thomas, Chachoua, Ilyas, Tisserand, Amandine, Vertenoeil, Gaëlle, Nédélec, Audrey, Vertommen, Didier, Roy, Anita, Marty, Caroline, Nivarthi, Harini, Defour, Jean-Philippe, El-Khoury, Mira, Hug, Eva, Majoros, Andrea, Xu, Erica, Zagrijtschuk, Oleh, Fertig, Tudor E., Marta, Daciana S., Gisslinger, Heinz, Gisslinger, Bettina, Schalling, Martin, Casetti, Ilaria, Rumi, Elisa, Pietra, Daniela, Cavalloni, Chiara, Arcaini, Luca, Cazzola, Mario, Komatsu, Norio, Kihara, Yoshihiko, Sunami, Yoshitaka, Edahiro, Yoko, Araki, Marito, Lesyk, Roman, Buxhofer-Ausch, Veronika, Heibl, Sonja, Pasquier, Florence, Havelange, Violaine, Plo, Isabelle, Vainchenker, William, Kralovics, Robert, Constantinescu, Stefan N.
• Format: Journal Article Web Resource
• Language: English
• Published: United States Elsevier Inc 23.02.2023; American Society of Hematology
• Subjects: Biochemistry; Calreticulin - genetics; Cell Biology; Cytokines - metabolism; Hematology; Human health sciences; Humans; Hématologie; Immunologic Factors; Immunology; Janus Kinase 2 - genetics; Life Sciences; Mutation; Myeloproliferative Disorders - genetics; Neoplasms; Sciences de la santé humaine
• ISSN: 0006-4971; 1528-0020; 1528-0020
• DOI: 10.1182/blood.2022016846

•Mutant CALR proteins are secreted in complex with sTFR1 to the plasma of MPN patients and activate the TpoR in a rogue cytokine fashion.•TpoR-expressing cells with a CALR mutation are uniquely sensitive to the levels of circulating mutant CALR proteins seen in patients. [Display omitted] Mutant calreticulin (CALR) proteins resulting from a −1/+2 frameshifting mutation of the CALR exon 9 carry a novel C-terminal amino acid sequence and drive the development of myeloproliferative neoplasms (MPNs). Mutant CALRs were shown to interact with and activate the thrombopoietin receptor (TpoR/MPL) in the same cell. We report that mutant CALR proteins are secreted and can be found in patient plasma at levels up to 160 ng/mL, with a mean of 25.64 ng/mL. Plasma mutant CALR is found in complex with soluble transferrin receptor 1 (sTFR1) that acts as a carrier protein and increases mutant CALR half-life. Recombinant mutant CALR proteins bound and activated the TpoR in cell lines and primary megakaryocytic progenitors from patients with mutated CALR in which they drive thrombopoietin-independent colony formation. Importantly, the CALR-sTFR1 complex remains functional for TpoR activation. By bioluminescence resonance energy transfer assay, we show that mutant CALR proteins produced in 1 cell can specifically interact in trans with the TpoR on a target cell. In comparison with cells that only carry TpoR, cells that carry both TpoR and mutant CALR are hypersensitive to exogenous mutant CALR proteins and respond to levels of mutant CALR proteins similar to those in patient plasma. This is consistent with CALR-mutated cells that expose TpoR carrying immature N-linked sugars at the cell surface. Thus, secreted mutant CALR proteins will act more specifically on the MPN clone. In conclusion, a chaperone, CALR, can turn into a rogue cytokine through somatic mutation of its encoding gene. Mutations in calreticulin (CALR), an endoplasmic reticulum chaperone, modify the protein’s C-terminus, leading to intracellular ligand-independent thrombopoietin receptor signaling and causing myeloproliferative neoplasms (MPNs). Mutant CALR is also found in a soluble form, but its significance is unknown. Pecquet et al revealed that soluble mutant CALR acts in a paracrine fashion to selectively enhance the growth of surrounding CALR-mutant hematopoietic cells. These novel insights suggest new therapeutic opportunities to target CALR-mutant MPNs.