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308 Part IV Disorders of Hematopoietic Cell Development

of the Friend erythroleukemia complex encodes the glycoprotein JAK2V617F in patients with Ph-negative myeloproliferative disorders
F-gp55, which binds and activates murine EPOR. 302–304 F-gp55 has been confirmed by numerous publications. Depending on the
appears to bind to EPOR via its transmembrane region. EPO and study, JAK2V617F has been reported in 60% to 90% of patients
F-gp55 binding sites are discrete; tertiary complexes of EPOR, EPO, with PV, 30% to 50% of patients with essential thrombocythemia
and F-gp55 have been detected on the surface of Friend virus-infected (ET), and 30% to 60% of patients with PMF. The mutation is
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cells. Second, EPOR can be constitutively activated by a point harbored at either the heterozygous or, by somatic recombination,
mutation (R129C) in the extracytoplasmic region of the polypep- the homozygous stage. It is detectable in all myeloid cells up to the
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tide. This mutation occurs in the “dimerization domain” of EPOR clonal hematopoietic stem cells. The mutation affects the domain
and results in constitutive homodimerization of the EPOR polypep- of the protein that is not directly involved in signal transduction but
tide, presumably through a disulfide bond. This mutation further is necessary to return the protein to its resting configuration after
underscores the importance of receptor dimerization in the initiation signaling. As a consequence, after its first engagement with EPO,
of a receptor signaling response. Third, EPOR can be constitutively EPOR signaling becomes constitutively activated. Because JAK2 is
activated in an autocrine manner. Murine erythroleukemia cell lines the earliest element of the EPOR pathway, it is conceivable that
have been established that coexpress EPOR and EPO. A fourth the high red cell numbers found in patients with PV carrying the
mechanism of EPOR constitutive activation results from EPOR JAK2V617F mutation is a direct consequence of constitutive EPO
overexpression. For instance, some murine erythroleukemia cell lines signaling in erythroid cells. The extent to which the downstream
have increased EPOR mRNA, resulting from spleen focus-forming EPOR signaling pathway is altered as a consequence of this constitu-
virus proviral integration within the first intron of the murine EPOR tive activation is a debatable issue. The observation that erythroblasts
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gene. Overexpression of the normal murine EPOR polypeptide derived in vitro from patients with PV carrying the JAK2V617F
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may thereby contribute to oncogenesis. mutation are highly resistant to TRAIL-induced apoptosis suggests
Soon after the cDNA of mouse and human EPOR were cloned, that the mutation exquisitely increases erythroblast survival. However,
the mouse and human genomic structures were identified. The gene additional abnormalities induced by the presence of JAK2V617F are
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for mouse EPOR was found to map to mouse chromosome 9, represented by hypersensitivity to insulin-like growth factor I and
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whereas the human gene was found on human chromosome 19p. increased EPOR recycling from the Golgi apparatus. More difficult
Mapping of the EPOR genes led to their implication in various to understand is why the mutation is also present in some patients
human disease states. For instance, studies demonstrated that a chro- with ET and PMF. JAK2, in addition to EPOR, has an important
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mosomal breakpoint 3′ to the human EPOR gene results in increased role in transducing the signal from c-Mpl, the TPO receptor. The
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EPOR expression. The rearranged EPOR allele appears to encode a protein has two opposing effects on the intracellular processing of
mutated EPOR polypeptide with increased activity, perhaps second- the two receptors after growth factor stimulation: it favors EPOR
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ary to loss of the C-terminal negative regulatory domain. recycling from the Golgi apparatus, but it determines retention and
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EPOR plays a role in the rare congenital disease familial degradation of c-Mpl in the cytoplasm. As such, the JAK2V617F
erythrocytosis. Familial erythrocytosis is a heterogeneous group of mutation should increase the number of EPOR on the surface of
hereditary conditions characterized by an increase in red blood cell erythroid cells while reducing that of c-Mpl on megakaryocytes. This
mass in the setting of low serum EPO levels. A few families that hypothesis explains why the mutation is found preferentially in PV.
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demonstrate autosomal dominant inheritance have been identified. However, megakaryocytes originating from JAK2V617F stem cells
The linkage between the EPOR gene and familial erythrocytosis might express a reduced number but constitutively active c-Mpl, so
was first established by the observation that a mutant EPOR allele it can be argued that the mutation manifests itself prevalently in the
segregates with the disease in one familial erythrocytosis kindred. erythroid or the megakaryocytic lineage, depending on genetic poly-
This allele contains a nonsense mutation in the coding region of morphisms outside the JAK2 locus present in the population. This
the gene that results in synthesis of a truncated EPOR that lacks the hypothesis is supported by the observation that mice transplanted
negative regulatory domain of its C-terminal region. 312,313 Since that with JAK2V617F hematopoietic stem cells develop either a PV- or
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report, several other frameshift and deletion EPOR gene mutations, an ET-like syndrome, depending on their genetic background and/
all encoding C-terminal–truncated forms of the protein providing or level of JAK2V617F expression. 332,333 One of the genetic factors
EPO hypersensitivity and resulting in familial erythrocytosis, have that may determine the phenotype expressed by JAK2V617F-positive
been reported. 314–318 myeloproliferative neoplasms may be represented by the GR locus.
Congenital polycythemia may arise not only from gene mutations The blood mononuclear cells from PV patients, but not those from
leading to abnormal EPOR signaling but also from gene mutations ET or PMF, express increased levels of the dominant negative GRβ
altering EPO production. The T598T mutation in one of the genes isoform. Increased levels of GRβ are also expressed by the erythro-
controlling oxygen sensoring (von Hippel-Lindau [VHL] gene), blasts expanded in vivo from PV patients. The observations that these
which leads to increased EPO production by the kidney, is associated erythroblasts lack nuclear STAT5 activity, in spite of constitutive
with congenital erythrocytosis, the Chuvash polycythemia. 319,320 The STAT5 phosphorylation induced by the presence of JAK2V617F
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C598T VHL mutation is endemic in Chuvashia, Russian Federation, mutation, and do not mature in response to EPO suggest that
and in the small island of Ischia, in southern Italy. The frequency constitutive inhibition of the EPO maturation signal provided by
of the mutation on this small island is higher than in Chuvashia GRβ expression may confer a self-renewal state to PV erythroblasts,
(0.07% versus 0.02%), but the haplotype of Italian patients matches contributing to erythrocytosis. Expression of GRβ may be favored
that identified in the Chuvash cluster, supporting a single-founder by the increased frequency of the rs6198 SNP, which stabilize GRβ
hypothesis. 321 mRNA observed in these patients. 124
Acquired mutations in the EPOR signaling pathway have Studies on the biology and biochemistry of the JAK2V617F muta-
been identified in PV, essential thrombocythemia, and idiopathic tion are actively being pursued by many investigators. Furthermore,
myelofibrosis, a class of myeloproliferative neoplasm that lacks the in JAK2V617F-negative PV or idiopathic erythrocytosis patients,
Philadelphia chromosome (Ph) abnormality and therefore cannot several other gain-of-function mutations affecting JAK2 exon 12 with
be cured with inhibitors such as Gleevec, which are specific for a distinct phenotype (idiopathic erythrocytosis) have been identi-
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the signaling pathway altered by the Ph abnormality. These diseases fied. The identification of specific mutations and the availability
originate at the level of the pluripotent hematopoietic stem cell and of the crystal structures of EPO, EPOR, and JAK2 has allowed
are characterized by proliferation of one or more of the myeloid computer modeling to design targeted protein-signaling inhibitors
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lineages but relatively normal hematopoietic cell maturation. Several for treatment of Ph-negative myeloproliferative neoplasms. In as
investigators have reported a mutation in the JAK2 gene result- few as 9 years after the discovery of JAK2 mutations, JAK2 inhibitors
ing in a valine→phenylalanine substitution at position 617 of the have been designed, clinical trials with selected drugs conducted,
protein (JAK2V617F mutation) in patients with Ph-negative chronic and the drugs approved for clinical use by the U.S. Food and Drug
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proliferative disorders. 322–325 Since the early reports, the presence of Administration. The observation that the genetic lesions found in