Mutant Calreticulin in MPN: Mechanistic Insights and Therapeutic Implications

Abstract

Purpose of Review

More than a decade following the discovery of Calreticulin (CALR) mutations as drivers of myeloproliferative neoplasms (MPN), advances in the understanding of CALR-mutant MPN continue to emerge. Here, we summarize recent advances in mehanistic understanding and in targeted therapies for CALR-mutant MPN.

Recent Findings

Structural insights revealed that the mutant CALR-MPL complex is a tetramer and the mutant CALR C-terminus is exposed on the cell surface. Targeting mutant CALR utilizing antibodies is the leading therapeutic approach, while mutant CALR-directed vaccines are also in early clinical trials. Additionally, chimeric antigen receptor (CAR) T-cells directed against mutant CALR are under evaluation in preclinical models. Approaches addressing the cellular effects of mutant CALR beyond MPL-JAK-STAT activation, such as targeting the unfolded protein response, proteasome, and N-glycosylation pathways, have been tested in preclinical models.

Summary

In CALR-mutant MPN, the path from discovery to mechanistic understanding to direct therapeutic targeting has advanced rapidly. The longer-term goal remains clonally-selective therapies that modify the disease course in patients.

Introduction

Myeloproliferative Neoplasms (MPN) are a group of rare chronic blood cancers with an incidence of approximately 4 in 100 000 people [1]. In the majority of cases, MPN are caused by one of three driver mutations in JAK2 [2–5], CALR [6, 7] or MPL [8] arising in the hematopoietic stem cell (HSC) compartment. These driver mutations, which are strongly mutually exclusive, lead to constitutively active signaling in the JAK-STAT pathway resulting in excessive proliferation of HSCs and expansion of differentiated myeloid cells in the peripheral blood, leading to three distinct disease subtypes: essential thrombocythemia (ET), polycythemia vera (PV) and primary myelofibrosis (PMF) [9, 10]. While most patients harbor a JAK2 mutation (50–60%), approximately 25–30% of all cases arise due to mutated CALR, which exclusively present as ET or PMF [6]. Due to their important role as disease-initiating mutations, CALR mutations in MPN have been the subject of intense research over the last decade since their discovery in 2013 [6, 7].

Wild-type CALR is a highly conserved chaperone protein residing in the endoplasmic reticulum (ER), overseeing correct protein folding [11] and regulating calcium storage [12]. MPN-associated mutations in CALR occur exclusively in the last exon [6, 7]. Although a variety of different CALR mutations can act as MPN driver mutations, the most frequent CALR mutations are a 52-base pair (bp) deletion (L367fs*46) (53%) and a 5-bp insertion (K385fs*47) (31.7%) [6, 7]. Irrespective of the specific DNA mutation, all CALR mutations result in a + 1 bp frameshift, leading to the generation of a common, shared C-terminal peptide [13]. This C-terminally altered mutant CALR binds to immature MPL in the ER, and the complex traffics to the cell surface, activating the JAK-STAT pathway [14–19].

In this review article, we focus on the most recent advances in mutant CALR biology and their relevance for targeting mutant CALR therapeutically.

A Pathogenic Binding Interaction Between Mutant CALR and MPL Drives MPN Development

Recurrent somatic mutations in CALR were first described in MPN in 2013 [6, 7] but the mechanism underlying the MPN phenotype was unclear. Early investigations revealed that mutant CALR is oncogenic only in the presence of thrombopoietin (TPO) receptor, MPL. Mutant CALR-expressing cells (Ba/F3 and UT-7) are transformed to cytokine-independence when MPL is co-expressed, but not in the presence of other type I cytokine receptors (i.e., EPOR or GCSF-R) [15–17]. Subsequently, it was shown that mutant CALR forms a homomultimeric complex that facilitates pathogenic binding to MPL [20]. Meanwhile, studies demonstrated that both N-glycosylation of MPL’s extracellular domain [14] and the lectin-binding sites of mutant CALR [14, 21] are essential for the pathogenic interaction between the two proteins and subsequent activation of oncogenic JAK-STAT signaling. Pecquet and colleagues demonstrated that mutant CALR binding to immature N-glycans on MPL facilitates trafficking of MPL to the cell surface as a stable complex with mutant CALR and that a hydrophobic patch in the extracellular domain of MPL is necessary for MPL activation by mutant CALR [22].

Recent work by Papadopoulos et al. revealed the binding kinetics between mutant CALR and MPL [23]. Utilizing hydrogen-deuterium exchange mass spectrometry, the authors demonstrated that the modified mutant C-terminus of CALRdel52 also affects the N-domain of CALR [23], and makes it more accessible to immature N-glycan binding on MPL (Fig. 1A). They further discovered that the mutant CALR-MPL complex is a tetramer, with mutant CALR dimers engaging with the extracellular domain of MPL at two key sites: the N-terminus of mutant CALR binding to immature N-glycans on MPL and the mutant C-terminus residues binding to acidic patches in MPL [23]. Importantly, these structural studies found that the mutant CALR C-terminus of the mutant CALR-MPL complex is exposed on the cell surface.

Fig. 1.

Therapeutic targeting in CALR-mutant MPN. A) The mutant CALR-MPL tetrameric complex on the cell surface, showing the mutant CALR C-terminus is exposed and that mutant CALR interacts with immature N-glycans in the extracellular domain of MPL to constitutively activate the JAK-STAT pathway. B) Treatment with pegylated IFNα generally leads to a significantly reduced variant allele frequency in JAK2V617F + MPN patients but not in CALR-mutant MPN patients. C) Mutant CALR-specific antibodies can bind to the mutant CALR-MPL complex on the cell surface to block MPL-JAK-STAT signaling. Addition of a JAK2 inhibitior (e.g. ruxolitinib) may further inhibit JAK-STAT signaling. D) Bi-specific antibodies can bind both mutant CALR bound to MPL, as well as the CD3 receptor, to bring CALR-mutant MPN cells and T-cells in close proximity to facilitate T-cell killing. E) Combining a mutant CALR-directed vaccine with a checkpoint inhibitor (e.g. ipilimumab), or an adjuvant (e.g. poly-ICLC and keyhole limpet hemocyanin), are therapeutic strategies under evaluation in early phase clinical trials

Understanding how mutant CALR binds and activates MPL is of fundamental importance to understanding the pathogenesis of CALR-mutant MPN as well as identifying potential vulnerabilities that could be pharmacologically exploited.

CALR-Mutant HSC Display a Megakaryocyte-Lineage Differentiation Bias and Molecular Resistance to Interferon Alpha (IFNα)

CALR mutations arise in the HSC compartment [6, 7] and are sufficient alone to cause MPN [24]. Using genotyping of transcriptomes (GoT), Nam et al. demonstrated that the CALR-mutant malignant clone represents a subpopulation of hematopoietic stem and progenitor cells (HSPCs) that coexist with CALR-wildtype cells [25]. The authors showed that the proportion of mutated cells is not equally distributed within the progenitors, with CALR-mutated HSPCs biased towards myeloid differentiation and CALR-mutated cells heavily enriched in megakaryocyte progenitors [25]. Using mathematical modeling, Hermange and colleagues concluded that the CALR-mutant malignant clone displays a proliferation and fitness advantage [26]. They further reported that while CALR mutations appear to be acquired later in life than JAK2^V617F, CALR-mutant HSCs expand faster compared to JAK2^V617F HSCs, suggesting a higher proliferative advantage for CALR-mutant HSC clones in the BM [26]. These findings have implications for therapeutic targeting.

Interferon alpha (IFNα) and its pharmaceutical derivatives have been used in the clinic as a cytoreductive therapy for MPN for more than 30 years, due to their ability to induce hematological responses in patients with PV and ET [27, 28]. Mechanistic studies in mice have shown that IFNα triggers the cell cycle entry of quiescent HSCs leading to HSC exhaustion [29, 30]. Czech et al. showed in their retrospective analysis of MPN patients treated with pegylated IFNα that despite having a similar level of clinic-hematological response, patients with CALR-mutated MPN had no significant reduction in variant allele frequency (VAF) compared to JAK2^V617F patients [31]. Similarly, a prospective longitudinal analysis of HSCs from 48 MPN patients reported that the CALR-mutant VAF remained stable upon IFNα treatment, while the JAK2^V617F VAF decreased over time due to IFNα-induced differentiation and depletion of JAK2-mutant HSCs (Fig. 1B) [32]. Molecular profiling of the DALIAH trial (NCT01387763), a prospective randomized trial of pegylated interferon or hydroxyurea, also found distinct molecular responses to pegylated interferon in JAK2- and CALR-mutant patients, with superior molecular responses in JAK2-mutant patients [33].

In aggregate, CALR-mutant HSC appear to expand rapidly in the bone marrow (BM) to cause MPN but despite this are not effectively targeting with pegylated interferon. This poses a therapeutic challenge, particularly for young patients for whom pegylated interferon is often a preferred treatment option. The mechanism underlying the apparent molecular resistance of CALR-mutant HSC to pegylated interferon is an area of active study. In parallel, novel approaches to target mutant CALR directly on the cell surface are currently under active investigation.

Therapeutically Targeting Mutant CALR on the Cell Surface

Given the role of aberrant mutant CALR-MPL signaling in disease pathogenesis and the accessibility of the complex on the cell surface, immunotherapeutic approaches have been investigated in preclinical models and a subset have advanced to the clinic (Fig. 1C).

B3, a mouse chimeric monoclonal antibody (mAb) that binds the mutant C-terminus of CALR was employed in an initial study by Kihara et al., presented at the 2020 American Society of Hematology meeting [34]. Their findings showed that intravenous injection of B3 reduced CALRdel52-induced thrombocytosis and decreased megakaryocytes in the BM of an ET mouse model, making it a potentially effective targeted treatment strategy [34]. Subsequently, Achytuni et al. reported that CALRdel52-bearing transgenic mice treated with a mouse IgG2a anti-human CALR-mutant mAb exhibited a reduction in platelets and lineage (Lin)⁻ Sca-1⁺ c-Kit⁺ (LSK) cells in the BM as compared to vehicle-treated mice [19]. One concern in this study was the rapid decrease in platelet levels in mice within six hours post-intraperitoneal injection with the CALR-mutant mAb, the cause for which remains unclear.

In 2022, Mughal et al. highlighted the potential of targeting CALR-mutant cells using antibodies raised against peptides within the mutant CALR C-terminus. The majority of antibodies they generated reacted to a specific epitope within the mutant CALR frameshift region, however functional studies in mutant CALR-expressing cells were not performed [35]. In the same year, Tvorogov et al. developed 4D7, a rat IgG2α mAb that binds to the mutant C-terminus of CALR and showed the 4D7 antibody distrupts CALR-MPL complex at the cell surface, thereby decreasing constitutive activation of JAK-STAT signaling [36]. The authors also found that 4D7 suppressed TPO-independent proliferation and megakaryocyte differentiation of patient-derived CALR-mutant CD34⁺ cells [36]. Furthermore, treatment with 4D7 increased the survival of CALR-mutant xenograft mouse models [36].

Recently, a significant breakthrough in targeting mutant CALR was achieved with the development of INCA033989, a fully human IgG1 mutant CALR mAb [37]. In vitro studies of INCA033989 in CALR-mutant Ba/F3-MPL and UT-7-MPL cells showed that the mAb is capable of binding to mutant CALR on the cell surface, blocking the mutant CALR-MPL interaction and inhibiting JAK-STAT signaling [37]. Treatment with INCA033989 inhibited JAK-STAT signaling in patient derived CD34⁺ cells while leaving signaling intact in CD34⁺ control cells from healthy donors, indicating the antibody’s specificity [37]. Additionally, in vivo experiments demonstrated that INCA033989 treatment in a mutant CALR conditional knockin mouse model prevented thrombocytosis and significantly decreased CALR-mutant HSCs in the BM [37]. Although the authors demonstrated that the mechanism of action of INCA033989 was partially mediated through dynamin-dependent endocytosis of the INCA033989/mutant CALR-MPL complex, this aspect of the study requires additional validation, particularly with respect to the effect of INCA033989 on the conformation of the mutant CALR-MPL complex and on MPL recycling. Interestingly, surface plasmon resonance (SPR) studies revealed that INCA033989 has preferential binding to recombinant CALRdel52 mutant protein as compared to CALRins5 mutant protein, suggesting that INCA033989 may have greater activity in MPN patients with a Type I CALR mutation as compared to patients with Type II mutation. INCA033989 has now entered a phase 1 clinical trial (NCT06034002), a non-randomized, open label multicenter study in patients with CALR-mutant MPN. The trial is currently recruiting, no results have yet been reported.

Another novel approach involves the T-cell redirecting bispecific antibody, JNJ-88549968, which targets mutant CALR on the cell surface and acts as a bridge between CALR-mutant cells and T-cells [38] (Fig. 1D). This enhances the accessibility of CALR-mutant expressing cells for T cell-mediated cytotoxicity and may make it more effective in eliminating CALR-mutant cells as compared to a Fc silent, blocking mutant CALR antibody. The authors reported concentration-dependent cytotoxicity of patient-derived CALR-mutant CD34 + cells upon treatment with JNJ-88549968 and demonstrated in vivo efficacy in patient-derived xenograft mouse models, prolonging the survival of treated mice compared to vehicle-treated controls [38]. A potential challenge with JNJ-88549968 is that this bispecific antibody necessitates a functional T-cell repertoire in CALR-mutant patients to effectively eliminate the CALR-mutant cells. This may pose a challenge, as some MPN patients may can lack functional cytotoxic T-cells due to the increased expression of checkpoint inhibitory receptors and T-cell exhaustion [39, 40]. Because of the promising results in the preclinical studies, JNJ-88549968 has also now entered a clinical trial (NCT06150157), in the form of a phase 1 open label interventional study that is presently recruiting participants.

While these studies advance targeted therapeutics tailored specifically for CALR-mutant MPN, a potential challenge to targeting mutant CALR on the cell surface using antibodies is soluble mutant CALR. In addition to cell surface translocation while bound to MPL, mutant CALR is also passively secreted and detectable in cultured supernatants [41–43], as well as in the plasma of MPN patients at levels of up to 160 ng/mL [44]. The presence of secreted mutant CALR could act as a decoy and prevent the mAbs from binding to MPL-bound mutant CALR. As data emerges from the ongoing phase 1 clinical trials, it will be interesting to see if comparable results are observed in patients, as compared to those reported in preclinical studies in mice.

Finally, CAR-T cell therapy, which involves engineering T-cells to express chimeric antigen receptors (CARs) that target specific proteins on the cell surface, may also hold promise for CALR-mutant MPN. At the 2024 European Hematology Association (EHA) meeting, Schueller et al. reported the ontarget efficacy of fully murine anti-mutant CALR CAR-T cells against Ba/F3-hMPL cells expressing human CALRdel52 in vitro as well as in vivo in transplanted NSG mice [45]. While this proves that CAR-T cells can effectively target primary CALR-mutant cells, in vivo treatment of immunocompetent chimeric mice did not result in disease clearance. More recently, Rampotas et al. report in their abstract, which will be presented at the 2024 ASH meeting, that human CAR-T cells eradicate CALR-mutant HSPCs in vitro as well as in vivo in immunodeficient mice [46]. This study also reports the potential of mutant CALR-directed CAR-T cell therapy to reduce mutant CALR allele burden and deplete CALR-mutant clones in the BM of xenograft mouse models. Additionally, they report that the efficiency of the CAR-T cells is lower in samples low in MPL receptor expression and that CAR-T cell killing is reduced in samples with prolonged exposure to JAK inhibitors, which could potentially pose a challenge in future clinical applications [46].

While CAR-T cells can be engineered to recognize unique epitopes in the mutant CALR C-terminus, challenges such as impaired functionality of T-cells in MPN patients and immune-related toxicity are likely to be relevant. Mutant CALR directed CAR-T cell therapy has not yet entered clinical trials.

Vaccination and T-Cell Directed Approaches to Target Mutant CALR C-Terminus

Another therapeutic approach that has advanced to the clinic is the development of a vaccine targeting the mutant C-terminus of mutant CALR.

Initial studies from Denmark focused on T-cells in patients with CALR-mutant MPN. Holmstrom et al. reported that T cells derived from patients with CALR-mutant MPN are reactive to mutant CALR C-terminus peptides following ex vivo stimulation and in response produce IFNγ [47]. In 2018, the same group reported that CD4⁺ T cells isolated from CALR-mutant MPN patients can recognize and kill autologous CALR-mutant cells ex vivo [48]. However, later, using peptides targeting the mutant CALR C-terminus, the same group found that while T cells from CALR-mutant MPN patients respond to peptide stimulation, they exhibited a decreased reactivity as compared to healthy donors stimulated with the same mutant CALR targeting peptides [49]. Additionally, the authors found that mutant CALR-reactive T cells in healthy individuals are CD4⁺ memory cells, raising the possibility that endogenous mutant CALR-directed immune responses may be an immunosurveillance mechanism that eliminates the early malignant clone before it manifests clinically [49]. These findings suggest that T cell-directed immunotherapeutic strategies could be another approach in the treatment of CALR-mutant MPN, however challenges such as T-cell exhaustion and T-cell tolerance, also exist.

To explore the question of whether mutant CALR neoepitopes are seen by CD8 + T-cells in humans, Gigoux et al. assessed MHC-I allele haplotypes in 308 MPN patients with JAK2- and CALR-mutant MPN [50]. They used an in silico-based bioinformatics approach to predict which mutant CALR C-terminus neo-epitopes bind to MHC-I alleles with high affinity [50]. They then compared the frequency of these MHC-I alleles in patients with JAK2- and CALR-mutant MPN and found that MHC-I alleles predicted to bind mutant CALR neo-epitopes with high affinity were de-enriched in CALR-mutant MPN patients as compared to JAK2-mutant patients i.e., evidence of MHC-I skewing. These findings provide indirect evidence that mutant CALR neo-epitopes are processed and presented by MHC-I, in the context of the appropriate MHC-I allele i.e., an allele that binds a mutant CALR neo-epitope with high affinity.

The first-in-human clinical trial using a mutant CALR peptide vaccine (NCT03566446) failed to show clinical or molecular responses, despite evidence of ex vivo T-cell responses following peptide stimulation in eight out of ten patients [51]. There are several potential reasons for the absence of clinical responses: (i) MHC-restriction i.e., the patient with CALR-mutant MPN may not possess an MHC-I allele that binds a mutant CALR neo-epitope with high affinity [50]. (ii) Impaired neo-epitope presentation: As a chaperone protein, CALR is essential for assembling MHC-I molecules with peptides. Arshad et al. showed that mutant CALR attenuates MHC-I assembly and protein-loading complex formation, which could impede antigen processing and presentation via MHC-I molecules [52]. (iii) Immune tolerance: In patients who have the appropriate MHC-I allele, the lack of response to peptide vaccination might be a consequence of immune tolerance in antigenspecific T cells, as a result of chronic antigen stimulation, including potentially by secreted mutant CALR in the plasma. (iv) Additional potential reasons include an immunosuppressive milieu in MPN patients, increased peripheral blood myeloid-derived suppressor cells (MDSCs) [53] and lack of adjuvant stimulation to the peptide vaccine.

Additionally, Bozkus et al. reported that administering programmed cell death protein 1 (PD1) and cytotoxic T lymphocyte antigen 4 (CTLA4) blocking antibodies to CALR-mutant MPN PBMCs resulted in the restoration of CALR-mutant specific T-cell responses, following peptide stimulation [39]. The authors also demonstrated, in a phase I/II clinical trial (NCT03065400), that pembrolizumab, a mAb against PD-1, significantly augmented the production of IFNγ and increased the frequency of peripheral CD4 + and CD8 + T cells in a patient harboring a type I CALR mutation [39]. However, despite evidence of increased T-cell frequency, no patients responded clinically in the pembrolizumab monotherapy trial [54]. A lack of functional T cells (i.e. T-cell exhaustion) and the predominance of JAK2^V617F mutations among the enrolled MF patients (i.e. absence of neo-epitopes in JAK2-mutant MPN) are potential reasons for the results.

Despite the unsatisfactory clinical outcomes of checkpoint inhibitor monotherapy trials, ongoing clinical studies are exploring immune activation in combination treatment modalities to enhance mutant CALR-directed T cell immune responses in patients (Fig. 1E). There are two studies of relevance: (i) NCT05444530: concurrent administration of VAC85135, a CALR-mutant neoantigen, with a CTLA4 monoclonal antibody (Ipilimumab), an immune checkpoint inhibitor and (ii) NCT05025488: mutant CALR peptide vaccine administration along with the immune modulators, keyhole limpet hemocyanin and polyinosinic: polycytidylic acid. The use of heteroclitic peptide-based vaccinations, where the peptide for vaccination is synthetically modified to bind to the MHC-I complex with high-affinity in order to elicit a robust immune response, has also been proposed as a strategy for increasing T cell-mediated immune responses in MPN patients [50]. This approach has been tested in preclinical studies only to date [50].

Cellular Consequences of Mutant CALR Expression Beyond MPL-JAK-STAT Pathway Activation

The cellular consequences of mutant CALR expression beyond MPL-JAK-STAT pathway activation remain incompletely understood. Given that exon 9 frameshift mutations may impair the chaperone activity of CALR [55], it is reasonable to hypothesize that cells expressing mutant CALR may undergo ER stress and consequently activate the unfolded protein response (UPR) through mechanisms that are either dependent or independent of the JAK-STAT pathway. Accordingly, studies have demonstrated that CALR-mutant cells experience UPR as a result of intra-cellular calcium imbalance and/or protein misfolding [55, 56].

Myeloperoxidase (MPO) expression has been found to be diminished in homozygous CALR-mutant MPN patients due to premature proteasomal degradation [57]. However, it is uncertain whether the MPO deficit in these patients is the direct result of a CALR chaperone malfunction. Building on the knowledge that CALR is involved in the folding of glycoproteins in the ER [58], a recent study utilizing limited proteolysis-coupled mass spectrometry has revealed profound alterations in protein abundance and structural protein changes in granulocytes of CALR-mutant MPN patients [59]. Notably, the study identified that glycoproteins and calcium-regulated proteins are predominantly affected. However, these proteomic pertubations were mainly observed in homozygous CALR-mutant granulocytes, whereas most patients are heterozygous for CALR mutations, leaving the implications of proteome alterations in these patients unclear. Further investigation into the proteome in HSPCs and megakaryocytes, rather than granulocytes, would be insightful, given that these are the primary cellular compartments affected in CALR-mutant MPN patients [60].

Studies by Nam et al. [25]. and Ibarra et al. [55]. have provided evidence that CALR mutations activate the UPR in megakaryocyte progenitors. Nam et al. found that genes involved in the UPR, (e.g. XBP1 and BiP), are transcriptionally upregulated in CALR-mutated cells [25]. Ibarra et al. reported that type I CALR-mutant cells, which lack C-terminal calcium binding sites, activate the IRE1α/XBP1 pathway more strongly than type II mutants [55]. Meanwhile, Jutzi et al. found that genes of the IRE1α/XBP1 branch of the UPR and proteasome pathways are enriched in the transcriptome of CALR-mutant murine HSCs as compared to WT HSCs [56]. The authors further showed that combined inhibition of the proteasome and IRE1α pathways preferentially reduced the abundance of CALR-mutant HSPCs over WT cells in a mouse model [56].

An unbiased approach using a whole-genome CRISPR knock-out depletion screen in CALRdel52 Ba/F3-MPL cells revealed the N-glycosylation pathway was differentially required in CALR-mutant transformed cells compared to control cells [61]. The authors then demonstrated a pharmacological vulnerability for CALRdel52-expressing Ba/F3-MPL cells to N-glycosylation inhibition with 2-deoxy-D-glucose (2-DG) [61]. Treatment with 2-DG preferentially targeted CALR-mutated over WT HPSCs in mice and reduced colony forming units–megakaryocytes (CFU-MK) in CALR-mutant patient BM samples as compared to healthy donor BM samples [61].

Collectively, the findings outlined in this section indicate that mutant CALR expression has additional cellular consequences beyond MPL-JAK-STAT activation. Understanding these could uncover new therapeutic avenues, including synthetic lethality approaches through combinatorial targeting with JAK-STAT dependent strategies. Despite the clinical efficacy of JAK1/JAK2 inhibitors, such as ruxolitinib in CALR-mutant PMF, a major limitation is the lack of clonal selectivity for CALR-mutant HSPCs. Whether combining JAK inhibition with mutant CALR-directed approaches (e.g. mAb) or with other indirect approaches to exploit mutant CALR vulnerabilities will enhance clonal selectivity in CALR-mutant patients remains to be determined.

Conclusion

The discovery of CALR mutations has fundamentally altered our understanding of the molecular pathogenesis of MPN and resulted in novel therapeutic approaches that have advanced rapidly to clinical testing in CALR-mutant MPN patients. Targeting mutant CALR on the cell surface with antibodies or cellular therapy is the leading strategy currently. Despite these exciting advances much remains to be understood, and it is likely that achieving the long-sought goal of clonally selective therapy in CALR-mutant MPN will require several iterations and refinements. The impact of concomitant mutations (i.e. MPN genomic complexity) on therapeutic response to mutant CALR-directed therapies remains to be determined. Nonetheless, mutant CALR remains a phenomenal therapeutic target in MPN, and the potential for disease-modification through early and effective targeting is a real possibility, one that the MPN field is eager to see realized.

Data Availability

No datasets were generated or analysed during the current study.