Essential thrombocythemia (ET) is a rare myeloproliferative neoplasm affecting 43.7 out of every 100,000 people in the United States. The disease is characterized by abnormally high platelet counts, mutational abnormalities in Janus Kinase 2 (JAK2)/Calreticulin (CALR)/myeloproliferative leukemia virus oncogene (MPL), and increased megakaryocyte production and differentiation. The average age of onset for patients with ET is between 65-70 years, but recent studies have demonstrated a downward trend in the age of diagnosis. Mechanistically, ET mutations cause the dimerization of JAK and upregulation of the JAK-STAT pathways. Common treatment approaches seek to use cytoreduction and platelet inhibition to lower the risk of a thrombotic event. Hydroxyurea and low-dose aspirin have been the gold-standard of treatment for ET patients. This thesis sought to compare the current available therapy with second-line treatments and investigational treatments. Anagrelide is a key second-line treatment for ET that is used in the event of intolerance to hydroxyurea. It acts through cytoreductive mechanisms which result in a decreased platelet count. Major bleeding is a severe adverse event associated with anagrelide. Interferons are another second-line defense in the treatment of ET despite a lack of FDA approval for this indication. Interferons act directly to reduce platelet counts and, unlike other drug classes, mount an immunological response against the JAK2 stem cells to reduce the allelic burden. An immunological approach to ET may be key to the sustained treatment of the disorder without a daily dosing regimen. Despite the promise of interferons, severe adverse effects limit the adherence of many patients to this class of drugs. JAK inhibitors are an investigational drug class that acts directly through the JAK-STAT pathway. JAK inhibitors have shown little efficacy in the treatment of ET and may be better suited for treatment in combination therapies. Telomerase inhibitors are one such investigational drug class that may pair well with JAK inhibitors for the treatment of ET. All of these drug classes were compared to hydroxyurea with respect to their pharmacokinetics, pharmacodynamics, and patient evaluation. Hydroxyurea and low-dose aspirin showed superiority in comparison to other drug classes due to their low toxicity profile and minimum adverse side-effects, high oral bioavailability and wide distribution, high adherence, and production of the most uniform response to reducing thrombotic events and platelet counts. The interferon drug class shows unique potential for the treatment of ET and should be placed above the second-line treatment standard of anagrelide due to its benefits in treatment of younger and pregnant patients. Interferons are the only class of drug for the treatment of ET that did not increase the risk of drug-related leukemogenic transformations. Despite non-adherence due to side-effects and lack of an oral administration, interferons are superior to anagrelide due to their longer dosing interval and immunological attack on JAK2 stem cells. Treatment of ET with anagrelide has shown similar efficacy to hydroxyurea and interferons in platelet reduction and rivals hydroxyurea in the prevention of thrombotic risk. Despite this benefit, the risk of bleeding associated with anagrelide is a significant disadvantage. Hydroxyurea and low-dose aspirin remain the current standard of treatment for patients with ET, although new approaches may soon be available.

The Mevalonate Pathway: A Potential Therapeutic Target for JAK2-driven Myeloproliferative Neoplasms Lori Nicole Griner Abstract Myeloproliferative neoplasms (MPNs) are diseases of hematopoietic stem cell origin and are characterized by uncontrolled growth of cells of the myeloid compartment. The Philadelphia chromosome negative classical MPNs, including polycythemia vera, essential thrombocythemia, and myelofibrosis, are diseases of dysregulated JAK2 signaling. In fact, the majority of MPN patients have activating mutations in JAK2 (e.g JAK2-V617F), a tyrosine kinase that contributes to the growth and survival of myeloid cells. While MPNs were first described over sixty years ago, a significant need remains to develop therapeutic strategies for them. Inhibitors of JAK2 are currently being developed, and one inhibitor, ruxolitinib, was recently approved for certain MPN patients. Ruxolitinib has made profound impacts on improving splenomegaly and constitutional symptoms in MPN patients, but it and other JAK2 inhibitors have not significantly reduced the JAK2 mutant allele burden, and thus such inhibitors have not induced remission in these patients. The current consensus in the MPN field supports JAK inhibition for the treatment of patients, but a further understanding of MPNs and JAK2 signaling, as well as improved JAK2 inhibitors, may be necessary for treating MPN patients. The work described in this dissertation has uncovered novel requirements for JAK2-V617F-driven signaling and transformation. We demonstrate that JAK2-V617F co-localizes with lipid rafts, cholesterol-rich microdomains within the plasma membrane that function to serve as platforms for signaling complex formation. Signaling complex formation is a necessary component for dysregulated signaling induced by JAK2-V617F. We provide evidence that cholesterol altering-lipid raft disrupting agents attenuate JAK2-V617F-driven signaling. We also show that cholesterol-lowering statins are effective at downregulating JAK2 signaling and inducing apoptosis in JAK2-V617F-driven cell lines. Importantly, we show that statins, inhibitors of the mevalonate pathway, inhibit the growth of primary MPN cells, while the same statin doses have no effect on healthy controls. Impressively, we demonstrate that statins cooperate with multiple JAK inhibitors, including ruxolitinib, to inhibit cell growth and induce apoptosis of JAK2-V617F-driven cells. This report establishes statin-mediated inhibition of the mevalonate pathway as a potential approach to improve MPN therapeutics. We propose future studies with statins and JAK2 inhibitors in the treatment of MPNs.