Ultraprecision Therapy for Type 1 vs Type 2 CALR+ MPN by Dual Epitope Targeting that Restores Ruxolitinib Sensitivity

Somatic frameshift mutations in the gene encoding calreticulin (CALR) give rise to myelofibrosis and are classified as Type 1 (del52) or Type 2 (ins5) according to the degree of wildtype sequence retained adjacent to the neopeptide, with each type conferring different clinical outcomes. Targeting strategies specific for Type 1 vs Type 2 mutations would have enormous clinical utility in the treatment and prevention of myelofibrosis as responses to tyrosine kinase inhibitors are not durable nor mutation-specific. Here we show that dual targeting of Type 1 (del52) mutant CALR with two monoclonal antibodies directed against distinct epitopes in CALR have significant advantages compared to single agent treatment in the eradication of primary megakaryocyte progenitors in vitro and in a humanized ossicle microenvironment leading to improved survival in xenograft models. Dual targeting was superior in blocking constitutive STAT5 and ERK phosphorylation induced by del52 and prevented accumulation of JAK2 phosphorylation, overcoming ruxolitinib resistance. In contrast, Type 2 mutations showed increased CALR dimerization and were partially resistant to antibody targeting but could be impacted by ruxolitinib triple combination. Together, our data demonstrate an ultra-precision medicine approach tailored to either Type 1 OR Type 2 mutation classes will be required for maximal efficacy and complete blockade of JAK/STAT signalling, with far-reaching implications for patient management.