Dysregulated JAK2 is at the apex of three classic oncogenic signaling cascades including STAT, PI3K/Akt/mTOR, and MAPK pathways. Their interrelationship and dependencies downstream of JAK2 in MPN cells is incompletely understood. Combined pathway inhibition in vitro might prove beneficial due to a synergistic impact on functionally redundant oncogenic axes, might allow lower doses of the different agents with better tolerability, and might avoid or delay the development of resistance.
Abbrevations: JAK: Janus Family of Kinases; STAT: Signal Transducer and Activator of Transcription Factors; MF: Myelofibrosis; MPN: Myeloproliferative Neoplasm; MDS: Myelodysplastic Syndrome; ET: Essential Thrombocythemia; BAT: Best Available Therapy; TNF: Tumor Necrosis Factor; IFN: Interferon; MAPK: Mitogen-Activated Protein Kinases; PI3K: Phosphatidylinositol-3’-Kinase; SOCS: Suppressor of Cytokine Signaling
The JAK-STAT pathway constitutes a signal transduction system through which a large spectrum of extracellular cytokines and nearly as many cognate transmembrane receptors converge towards an intracellular code employing four JAK kinases (JAK1, JAK2, JAK3, and TYK2) and seven STAT factors (STAT-1, -2, -3, -4, -5a, -5b and -6) . The activated STATs dimerize and translocate into the cell nucleus to influence DNA transcription, and regulating gene expression. The JAK-STAT pathways activate or suppress the transcription of a wide array of genes that affect cell growth and apoptosis such as SOCS, Nmi, Bcl-XL, p21, MYC, and NOS2 . Each member of the JAK family has a primary role in mediating a signaling process with some overlap between them. JAK2 is important for hematopoietic growth factors signaling such as Epo, GM-CSF, thrombopoietin, IL-3, IL-5, growth hormone and prolactin-mediated signaling .
Hyperactive JAK signaling is found in MPNs such as polycythemia vera, ET, and MF, 7.8% to 13% of chronic myelomonocytic leukemia and 1% to 4.2% of MDS. JAK2 (V617F)–positive AML is mostly observed in patients with a previous MPN, but cases of V617F in de novo AML have been
described. A gain-of-function mutation in the kinase domain, T875N, was reported in acute megakaryoblastic leukemia. JAK2 mutations are relatively rare in lymphoid malignancies except for ALL, most commonly in Down syndrome–associated ALL. JAK1, JAK2, and JAK3 mutations and translocations are observed in high-risk BCR-ABL B-cell ALL. Amplification of the JAK2 locus has been detected in Hodgkin lymphoma and mediastinal B-cell lymphoma. JAK2 mutations and translocations are very rare in solid cancers .
Type I JAK inhibitors precisely target the well-conserved ATP-binding pocket of JAK1 and JAK2 in both active and inactive conformations. Ruxolitinib (and other JAK inhibitors under clinical evaluation) falls within the type I family  which has insufficient efficacy and limited JAK inhibition potential . Their lack of selectivity implies that the tolerable doses lie within a narrow therapeutic window because of the collateral inhibition of other JAK members. Specific inhibitors for one of the four closely related JAK kinases could partially tackle this drawback because of the marked similarity in the active site within the family. Inhibitors that have a type II binding-mode specifically engage and stabilize inactive kinases by exploiting an additional, less conserved allosteric site directly adjacent to the ATP binding pocket, providing another handle for tuning
selectivity. Alternatively, a covalent inhibitor exploiting a nonconserved
cysteine residue in the active site of JAK3 has been
shown to have strong target specificity .
Myelofibrosis: Ruxolitinib is the only JAK inhibitor approved
for treatment of patients with MF. The drug mainly inhibits
dysregulated JAK-STAT signaling, present in all MF patients
irrespective of their JAK2 mutational status. It is not selective
for the mutated JAK2, which explains its efficacy in both JAK2-
positive and JAK2-negative MF . Of note, clinical responses in
MF patients were linked to suppression of increased serum levels
of proinflammatory cytokines such as IL-1, IL-6, TNF-α and IFN-γ
. Ruxolitinib has become the standard of care for intermediate
or high-risk patients with MF; it provides rapid and sustained
reduction of splenomegaly and improvement in symptoms and
quality of life, possibly resulting in prolongation of survival .
Ruxolitinib treatment failure in MF manifests as primary
refractoriness or secondary resistance. Primary refractoriness
can be defined as no or only minimal clinical response (<35%
reduction of spleen volume compared to baseline), whereas
secondary resistance is indicated by the loss of a previously
confirmed clinical response (such as splenic relapse) or
progression to leukemia . Long-term follow-up showed
that about half the patients enrolled in the COMFORT-I and
COMFORT-II studies discontinued treatment by 3 years, largely
because of loss of response and/or disease progression .
Primary refractoriness is rare (<10%) . The mechanisms
of resistance and/or loss of response to ruxolitinib have not
yet been deciphered . It could be expected that resistant
patients would have acquired mutations at the drug-binding
site of the target kinase . Acquisition of mutations in the
predicted ruxolitinib-binding region conferred resistance to
JAK inhibitors, have not been documented in patients . A
significant correlation between resistance and the absence of
mutations in JAK2 (V617F), MPL, TET2, and SRSF2 at diagnosis
has been identified .
The phenomenon of persistence to a JAK2 inhibitor (i.e., the
fact that JAK2 (V617F) mutated cells survive despite chronic
JAK inhibition) was ascribed to heterodimerization between
activated JAK2 and other members of the JAK family and was
shown to occur in patients treated with JAK2 inhibitors; however,
the clinical relevance of such observations remains unsettled .
There are several limitations to the use of JAK2 inhibitors in
MF patients. First, the safety and clinical efficacy of ruxolitinib
are unclear in patients present with platelet counts between
50×109/l to 100×109/l or even <50×109/l, and in patients
present with transfusion-dependent anemia, since ruxolitinib
have platelet lowering properties and can lead to anemia in
some patients. Second, it does not cure the disease and requires
continuing therapy to maintain response . Besides, there is no
biological evidence of a possible favorable effect of ruxolitinib on
the survival of MF patients, such as the achievement of a complete
or a partial remission, cytogenetic or molecular response, or
reversal of the bone marrow fibrosis . Lastly, the response
may be short lived in some patients and there are reports of
adverse events occurring at the time of withdrawal .
Ruxolitinib is also approved for the second-line treatment
of polycythemia vera and is being developed for ET . Patients
with ET who are refractory to or intolerant of hydroxyurea can
achieve clinically meaningful and durable reductions in platelet
and WBC counts and improvements in ET-related symptoms with
ruxolitinib treatment . However, ruxolitinib did not improve
treatment efficacy for most clinically relevant events compared
with BAT and was more frequently associated with anemia and
infections than was BAT .
Ruxolitinib is approved for MDS and is effective in reducing
splenomegaly. The medication is off-label for hypereosinophilic
syndrome, a report demonstrates successful treatment of
HES patients, who are intolerant to other treatments, with
ruxolitinib . The efficacy of ruxolitinib in GVHD was reported
in a murine model, and its potent activity was demonstrated
in 6 patients with corticosteroid-refractory GVHD. Findings
demonstrate that targeting JAK1/2 signaling in alloreactive T
cells is a powerful approach to GVHD inhibition. The key event
is inhibition of STAT3 phosphorylation by ruxolitinib during
an allogeneic immune response. In addition to, the drugspecific
effects on Treg development. Unlike, most conventional
immunosuppressive agents which target T-cell function,
ruxolitinib impair differentiation, maturation, and cytokine
production of dendritic cells, which may further increase its
efficacy in GVHD. Suppression of proinflammatory cytokines
such as IL-1b, IL-6, or IFN-γ which are considered hallmarks of
GVHD could potentially reduce disease severity .
Other JAK inhibitors with the potential for significantly less
myelosuppression or even improvement of anemia continue
to be tested . Type II JAK inhibition such as BBT594 can
overcome persistence to ruxolitinib in JAK2 (V617F)- and MPL
W515L–positive cells. In contrast with type I JAK inhibition,
inactive JAK2 is stabilized and activation loop phosphorylation
is decreased .
Several alternative options to target MPNs are being explored.
HSP90 inhibitors such as PU-H71 and AUY922 efficiently
impair JAK-mediated signaling and overcome resistance due to
secondary mutations. Histone deacetylase inhibitors, including
panobinostat, reduce JAK2 expression most probably due to effects on JAK2 mRNA expression, and increased proteasomal
JAK2 degradation. In vitro studies suggest synergistic effects
of PI3K/mTOR inhibitors or of the MEK1/2 inhibitor with JAK2
inhibitors. Combined pathway inhibition therapy might prove
beneficial due to a synergistic impact on functionally redundant
oncogenic axes, might allow for lower doses of the different
agents with better tolerability, and might avoid or delay the
development of resistance .