Stem Cell Involvement in Myeloproliferative Neoplasms

The clonal origin of myeloproliferative neoplasms (MPN) has been first implied by work of John W. Adamson, Philip J. Fialkov and co lleagues (1), who in 1976 performed X-chromosome inactivation pattern (XCIP) studies in patients with MPN using restriction fragment length polymorphisms in the X-chromosomal gene glucose-6-phosphate dehydrogenase (G6PDH) (2). They demonstrated that peripheral blood cells in two female patients with polycythemia vera (PV) expressed the G6PDH derived solely from one of the two parental X chromosomes, indicating that hematopoiesis is clonal (1). Later the same authors showed that lymphoid cells can also be part of the MPN clone in essential thrombocythemia (ET) and PV, implying that the MPN disease initiated at the level of a multipotent hematopoietic stem cell (HSC) (3). For many decades XCIP was the only methodology to study clonality in MPN. The major advantage of XCIP is that the analysis can be applied in the absence of any knowledge about the molecular and genetic alterations. Among the disadvantages are that only female patients can be studied and that the clone must have expanded reached clonal dominance. The latter has to do with the fact that the ratio in the inactivation of the two X-chromosomes must substantially deviate from the expected 50:50 random distribution, a phenomenon also called after its author Mary F. Lyon “Lyonization” (4), in order to reach a minimal threshold for statistical significance. In most cases this cutoff is set at 75:25 or 80:20 (5). Thus, XCIP is relatively insensitive and clonal dominance must reach 75–80 % in order to be detectable. The methodology to detect XCIP has progressed and many polymorphisms in X-chromosomal genes have been described that allow finding informative markers in most female patients (6). Some differences exist between assays that measure methylation of female X-chromosomes at the DNA level and methods that rely on the expression of mRNA from genes located on the X-chromosome. The latter methodology is preferable and yields more reliable results (7). Using XCIP it has been demonstrated that PV and primary myelofibrosis (PMF) are invariantly clonal, while a subgroup of ET patients has been reported to display polyclonal hematopoiesis (8, 9). Cytogenetic analysis provided additional tools for studying clonal hematopoiesis in MPN. However, only 10–15 % of PV patients have abnormal karyotype at diagnosis and the most common abnormalities include trisomies (+8, +9, +1), and deletions on chromosome 20q (del20q) (10–12). More recently, microarray analysis of single nucleotide polymorphisms (SNPs) and copy number variation have yielded deeper insights into the molecular pathogenesis of MPN and have helped identifying new genes mutated in MPN (13–15). From today’s perspective, the term “polyclonal ET”, which was established based on XCIP studies can be deceptive and has caused considerable confusion in the field. It is important to realize that the absence of clonality by XCIP simply indicates that there is no clone that has expanded and reached 75–80 % of the cells that were analyzed. However, due to the low sensitivity of XCIP, the presence of clones that have not yet reached clonal dominance cannot be excluded. Indeed, today we have an increasing number of somatic mutations that can be used as markers to identify clones within a mixture of hematopoietic cells (16). The JAK2V617F mutation activates the tyrosine kinase domain of JAK2 Stem Cell Involvement in Myeloproliferative Neoplasms