Myeloproliferative neoplasms (MPNs) are clonal hematologic malignancies typified by genetic alterations that activate JAK-STAT signaling. Chronic MPNs exhibit a propensity for transformation to secondary acute myeloid leukemia (sAML), which carries a dismal prognosis. The overall objective of our work is to better understand the pathogenetic mechanisms responsible for the initiation and development of MPNs, as well as the factors that drive evolution to sAML. Our laboratory research is broadly organized into two areas:
Dysregulated Signaling Networks in MPNs
The identification of the JAK2 V617F mutation in MPNs has led to the development of targeted inhibitors of JAK2, which provide significant symptomatic benefit but do not appear to be capable of eradicating the underlying malignant clone. These findings suggest that other signaling pathways may be dysregulated in a manner that allows survival of the malignant clone, even when JAK2 is inhibited. We seek to identify and evaluate these altered signaling pathways as potential targets for therapeutic intervention. Mass cytometry is a novel technology that merges aspects of flow cytometry with mass spectrometry and enables the simultaneous measurement of 30+ parameters at the single cell level (Fig 1). This approach enables the characterization of aberrant signaling across the entire hematopoietic continuum. Our ongoing work with this approach has revealed an extensive network of hyperactivated signaling in MPN stem/progenitor cells. A major focus of these efforts is to understand the in vivo effects of JAK2 inhibition in primary MPN cells. Ultimately, we aim to integrate data from these experiments with concomitant genetic analysis such that phenotype can be connected to underlying genotype.
Figure 1. Inherent spectral limitations of fluorescence cytometry can be overcome with elemental mass cytometry. Conjugating antibodies to elemental mass tags enables the simultaneous measurement of 30+ parameters using a CyTOF mass cytometer.
Genetic Complexity and Clonal Evolution in MPNs
The JAK2 V617F mutation is present in the majority of MPNs, and recent studies have identified recurrent mutations in several other genes including CALR. However, the full spectrum of genetic changes in MPNs, particularly in relationship to clonal evolution and disease evolution, remains incompletely understood (Fig 2). We are utilizing next-generation sequencing (NGS) technologies to delineate clonal hierarchy in MPNs. One project focused on the analysis of serial matched samples from a patient with a chronic MPN transformed to sAML. Whole genome sequencing (WGS) and capture-based deep sequencing validation was utilized to recapitulate clonal architecture accompanying disease evolution. In a similar fashion, ongoing work is focused on understanding how specific genetic alterations may predict response to targeted therapies (e.g. JAK inhibitors), and how clonal dynamics may be modulated by these newer therapeutic modalities. We are also utilizing patient-derived induced pluripotent stem cells (iPSCs) and CRISPR site-directed gene editing to interrogate the role of JAK2 V617F and other driver mutations in the clonal progression of MPNs.
Figure 2. Genetic complexity and clonal evolution in myeloproliferative neoplasms (MPNs). The initiation, development, and progression of MPNs are driven by the acquisition of specific genetic alterations over time. The complex relationship between different combinations of mutations and clonal dynamics related to these genetic changes are not fully understood.