000166004 001__ 166004
000166004 005__ 20260116163027.0
000166004 0247_ $$2doi$$a10.1136/jitc-2025-012706
000166004 0248_ $$2sideral$$a147396
000166004 037__ $$aART-2026-147396
000166004 041__ $$aeng
000166004 100__ $$aPesini, Cecilia
000166004 245__ $$aMutant calreticulin enables potent and selective CAR-T cell therapy in preclinical models of myeloproliferative neoplasms
000166004 260__ $$c2026
000166004 5060_ $$aAccess copy available to the general public$$fUnrestricted
000166004 5203_ $$aBackground The adoptive transfer of T cells engineered to express chimeric antigen receptors (CAR-T) has shown high efficacy and safety in treating various hematologic malignancies. However, many hematologic disorders, such as BCR::ABL1-negative myeloproliferative neoplasms (MPNs), lack effective treatment options. Some of these neoplasms are marked by a recurrent mutation that results in the expression of mutant calreticulin (mCALR), a neoantigen absent in healthy tissues, making it a highly specific and appealing target for CAR-T cell therapy. Methods Five distinct CARs were designed based on available monoclonal antibody sequences that target mCALR and were subsequently used to generate CAR-T cells. The most effective construct was selected through functional in vitro assays against mCALR-positive cell lines. Its efficacy was then evaluated in cell lines, patient-derived cells, and orthotopic xenograft models, assessing tumor burden, CAR-T cell infiltration, and animal survival. Bulk and single-cell RNA sequencing were performed on patient-derived cells and residual tumor cells from CART-treated mice, respectively, to investigate potential resistance mechanisms. The impact of the most relevant pathway alteration on CAR-T efficacy was also analyzed. Pharmacological rescue assays using targeted agents were then conducted. Results Among the five constructs, one demonstrated superior and specific cytotoxicity against mCALR-expressing cells, with no activity against mCALR-negative controls. This CAR-T cell also eliminated patient-derived MPN cells and controlled disease progression in xenograft models, which correlated with the persistence of CAR-T cells and tumor infiltration. Transcriptomic profiling of patient samples and residual tumor cells in spleens of treated mice revealed upregulation of anti-apoptotic proteins. Functional assays confirmed reduced CAR-T efficacy in Bcl-2 high cells, which was restored by co-treatment with venetoclax, indicating a viable combination approach to overcome resistance. Conclusions This study demonstrates, for the first time, the successful targeting of mCALR with CAR-T cells as a therapeutic strategy for MPNs. The chosen construct shows strong preclinical efficacy against established cell lines and patient-derived cells. Additionally, transcriptomic profiling uncovered apoptosis resistance mechanisms and supports a combination strategy with BH3 mimetics, such as venetoclax. These findings provide a compelling rationale for ongoing preclinical development and future clinical application of anti-mCALR CAR-T cells for the treatment of MPNs.
000166004 536__ $$9info:eu-repo/grantAgreement/ES/AEI/PID2020-113963RB-I00$$9info:eu-repo/grantAgreement/ES/AEI/PID2024-157582OB-I00$$9info:eu-repo/grantAgreement/ES/AEI/RYC2022-036627-I$$9info:eu-repo/grantAgreement/ES/DGA/B29-23R$$9info:eu-repo/grantAgreement/ES/ISCIII/B61-24$$9info:eu-repo/grantAgreement/ES/DGA/LMP139-21$$9info:eu-repo/grantAgreement/EC/H2020/101018587/EU/Individual and Collective Migration of the Immune Cellular System/ICoMICS$$9This project has received funding from the European Union’s Horizon 2020 research and innovation program under grant agreement No H2020 101018587-ICoMICS$$9info:eu-repo/grantAgreement/ES/ISCIII/CB21-13-00087$$9info:eu-repo/grantAgreement/ES/ISCIII/PI25-00236$$9info:eu-repo/grantAgreement/ES/ISCIII/RD24-0014-0015$$9info:eu-repo/grantAgreement/ES/MICINN/PID2022-136554OA-I00
000166004 540__ $$9info:eu-repo/semantics/openAccess$$aby-nc$$uhttps://creativecommons.org/licenses/by-nc/4.0/deed.es
000166004 655_4 $$ainfo:eu-repo/semantics/article$$vinfo:eu-repo/semantics/publishedVersion
000166004 700__ $$aGil-Bellido, Mario
000166004 700__ $$aMillán, Lorena S.
000166004 700__ $$aOñate, Carmen
000166004 700__ $$aCalvo-Pérez, Adanays
000166004 700__ $$0(orcid)0000-0002-1861-5981$$aSantiago, Llipsy
000166004 700__ $$aIglesias, Eldris
000166004 700__ $$aBernal, Jorge Paúl
000166004 700__ $$aAraujo-Voces, Miguel
000166004 700__ $$0(orcid)0000-0001-6139-5905$$aPaz Artigas, Laura$$uUniversidad de Zaragoza
000166004 700__ $$aGarcía-Martínez, Laura
000166004 700__ $$aRoig, Francisco J
000166004 700__ $$0(orcid)0000-0002-0163-8378$$aMovilla Meno, Nieves$$uUniversidad de Zaragoza
000166004 700__ $$0(orcid)0000-0002-9864-7683$$aGarcia-Aznar, José Manuel$$uUniversidad de Zaragoza
000166004 700__ $$aMenéndez-Jandula, Bárbara
000166004 700__ $$0(orcid)0000-0002-6326-5992$$aOlave, María Teresa$$uUniversidad de Zaragoza
000166004 700__ $$0(orcid)0000-0001-5068-7355$$aAzaceta Reinares, Gemma$$uUniversidad de Zaragoza
000166004 700__ $$aGarrote, Marta
000166004 700__ $$aAlvarez-Larrán, Alberto
000166004 700__ $$aGálvez, Eva M
000166004 700__ $$aSánchez Martínez, Diego
000166004 700__ $$0(orcid)0000-0002-9730-2210$$aArias, Maykel A$$uUniversidad de Zaragoza
000166004 700__ $$0(orcid)0000-0003-0154-0730$$aPardo, Julian$$uUniversidad de Zaragoza
000166004 700__ $$0(orcid)0000-0002-3888-7036$$aRamirez-Labrada, Ariel$$uUniversidad de Zaragoza
000166004 7102_ $$11003$$2443$$aUniversidad de Zaragoza$$bDpto. Anatom.Histolog.Humanas$$cArea Histología
000166004 7102_ $$11007$$2610$$aUniversidad de Zaragoza$$bDpto. Medicina, Psiqu. y Derm.$$cArea Medicina
000166004 7102_ $$11011$$2566$$aUniversidad de Zaragoza$$bDpto. Microb.Ped.Radio.Sal.Pú.$$cÁrea Inmunología
000166004 7102_ $$15004$$2605$$aUniversidad de Zaragoza$$bDpto. Ingeniería Mecánica$$cÁrea Mec.Med.Cont. y Teor.Est.
000166004 7102_ $$11002$$2050$$aUniversidad de Zaragoza$$bDpto. Bioq.Biolog.Mol. Celular$$cÁrea Biología Celular
000166004 773__ $$g14, 1 (2026), e012706 [17 pp.]$$pJ. immunotherap. cancer$$tJournal for immunotherapy of cancer$$x2051-1426
000166004 8564_ $$s7895756$$uhttps://zaguan.unizar.es/record/166004/files/texto_completo.pdf$$yVersión publicada
000166004 8564_ $$s3285982$$uhttps://zaguan.unizar.es/record/166004/files/texto_completo.jpg?subformat=icon$$xicon$$yVersión publicada
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000166004 951__ $$a2026-01-16-14:54:43
000166004 980__ $$aARTICLE