Impact of Molecular Genetics on Outcome in Myelofibrosis Patients after Allogeneic Stem Cell Transplantation.

Molecular genetics may influence outcome for patients with myelofibrosis. To determine the impact of molecular genetics on outcome after allogeneic stem cell transplantation, we screened 169 patients with primary myelofibrosis (n = 110), post-essential thrombocythemia/polycythemia vera myelofibrosis (n = 46), and myelofibrosis in transformation (n = 13) for mutations in 16 frequently mutated genes. The most frequent mutation was JAK2V617F (n = 101), followed by ASXL1 (n = 49), calreticulin (n = 34), SRSF2 (n = 16), TET2 (n = 10), U2AF1 (n = 11), EZH2 (n = 7), MPL (n = 6), IDH2 (n = 5), IDH1 (n = 4), and CBL (n = 1). The cumulative incidence of nonrelapse mortality (NRM) at 1 year was 21% and of relapse at 5 years 25%. The 5-year rates progression-free (PFS) and overall survival (OS) were and 56%, respectively. In a multivariate analysis CALR mutation was an independent factor for lower NRM (HR, .415; P = .05), improved PFS (HR, .393; P = .01), and OS (HR, .448; P = .03). ASXL1 and IDH2 mutations were independent risk factors for lower PFS (HR, 1.53 [P = .008], and HR, 5.451 [P = .002], respectively), whereas no impact was observed for "triple negative" patients. Molecular genetics, especially CALR, IDH2, and ASXL1 mutations, may thus be useful to predict outcome independently from known clinical risk factors after allogeneic stem cell transplantation for myelofibrosis.

Serial 18F-FDG PET for Monitoring Treatment Response After Allogeneic Stem Cell Transplantation for Myelofibrosis.

Our objective was to assess the feasibility of 18F-FDG PET/CT for noninvasive monitoring of treatment response after allogeneic stem cell transplantation (SCT) for myelofibrosis. METHODS: Twelve patients with myelofibrosis underwent 18F-FDG PET/CT before and after SCT. Bone marrow uptake, spleen uptake, and spleen size were assessed before and after SCT and compared with hematologic response criteria and bone marrow biopsies. RESULTS: All patients who did not achieve complete remission remained PET-positive (P = 0.02). Extent of disease, bone marrow metabolism, spleen metabolism, and spleen volume decreased significantly in patients with complete remission (P = 0.03). PET/CT after SCT had a sensitivity of 1.0 (95% confidence interval [CI], 0.54-1.0), a specificity of 0.83 (95% CI, 0.36-1.0), a negative predictive value of 1.0 (95% CI, 0.48-1.0), and a positive predictive value of 0.86 (95% CI, 0.42-1.0) for diagnosis of residual disease. CONCLUSION: 18F-FDG PET/CT is feasible for noninvasive monitoring of treatment response after allogeneic SCT for myelofibrosis.

Outcome after Transplantation According to Reduced-Intensity Conditioning Regimen in Patients Undergoing Transplantation for Myelofibrosis.

Allogeneic hematopoietic stem cell transplantation remains the sole curative option for myelofibrosis. Many transplantation recipients receive a reduced-intensity conditioning (RIC) regimen owing to age or comorbidities; however, there is little published evidence to guide the choice of RIC regimen. In this study, we compared outcomes in patients who received 1 of 2 frequently used RIC regimens for patients with myelofibrosis: fludarabine-busulfan (FB) and fludarabine-melphalan (FM). A total of 160 patients underwent a RIC allograft procedure (FB group, n = 105; FM group, n = 55). We have developed a complex statistical model involving weighting and adjustment to permit comparison between these 2 groups. After weighting, the incidence of acute graft-versus-host disease (GVHD) was 62% in the FM group and 31% in the FB group (P = .001), and the corresponding incidence of chronic GVHD was 49% and 53%, respectively. The 7-year progression-free survival was were 52% in the FM group versus 33% in the FB group, and the 7-year overall survival rate 52% in the FM group versus 59% in the FB group. Nonrelapse mortality (NRM) was 43% in the FM group and 31% in the FB group. Multivariable analyses revealed no significant differences in PFS between the 2 groups; however, the relapse rate was significantly lower in the FM group (hazard ratio, 9.21; P = .008), whereas a trend toward reduced NRM was seen in the FB group (hazard ratio, 0.51; P = .068). In conclusion, both regimens appear to be efficient in mediating disease control and can be used to successfully condition patients with myelofibrosis. The FM regimen appears to induce more NRM than the FB regimen, but with augmented control of disease, leading to comparable overall survival rates for both regimens.

Cytokine Expression Pattern in Bone Marrow Microenvironment after Allogeneic Stem Cell Transplantation in Primary Myelofibrosis.

The only curative therapy for primary myelofibrosis (PMF) is allogeneic stem cell transplantation (ASCT). However, although we know that patients can benefit from ASCT, we do not know the extent of the changes of the expression profile of cytokines and matrix modulation factors. In this first systematic analysis, we evaluated the expression profile of 103 factors before and after transplantation to identify potential biomarkers. The expression of fibrosis-, inflammation-, and angiogenesis-associated genes was analyzed in a total of 52 bone marrow biopsies: PMF patients (n = 14) before and after ASCT and, for control purposes, post-ASCT multiple myeloma patients (n = 14) and non-neoplastic hematopoiesis (n = 10). In post-ASCT PMF cases, decreased expression of tissue inhibitor of metalloproteinases (TIMP) and platelet-derived growth factor alpha (PDGFA) correlated with bone marrow remodeling and hematological remission. Expression of several other matrix factors remained at high levels and may contribute to post-ASCT remodeling. This is the first systematic analysis of cytokine expression in post-ASCT PMF bone marrow that shows that normalization of bone marrow microenvironment is paralleled by decreased expression of TIMP and PDGFA.

Prognostic factors for survival of patients with newly diagnosed chronic GVHD according to NIH criteria.

Chronic graft versus host disease (cGvHD) is the most common cause of late morbidity and mortality after allogeneic hematopoietic stem cell transplantation (HSCT). We retrospectively evaluated the impact of NIH classification on outcome of patients at our center. Primary endpoint was overall survival at 5 years. Two hundred one patients with cGVHD according to NIH were included. Platelets <100,000/µl on day of diagnosis of cGvHD (HR 2.97, 95 % CI 1.7-5.3, p < 0.001), female donor (HR 1.78, 95 % CI 1.0-3.2, p = 0.05), and reduced intensity conditioning (HR 1.95, 95 % CI 1.0-3.8, p = 0.05) impacted overall survival. Non-relapse mortality (NRM) was higher for patients with low vs. high platelets: 26 % (95 % CI 14-40) vs. 6 % (95 % CI 2-10), p < 0.001, and tended to be higher for female vs. male donor: 14 % (95 % CI 7-23) vs. 7 % (95 % CI 3-13), p = 0.08. Relapse tended to be higher for recipients of reduced intensity conditioning (RIC) vs. myeloablative conditioning (MAC): 33 % (95 % CI 23-43) vs. 20 % (95 % CI 10-31), p = 0.06. After excluding patients with myeloma and lymphoma, IgG serum levels at diagnosis of cGvHD of 122 patients were correlated with survival. IgG levels above normal were associated with worse 2-year overall survival (OS), p = 0.04, compared to normal or low IgG levels. Platelet count at diagnosis remains the most valid prognostic factor for survival of patients with cGvHD even in the era of NIH grading. High IgG level at diagnosis of cGVHD represents a potential negative prognostic parameter that deserves further investigation.

JAK1/2 inhibition impairs T cell function in vitro and in patients with myeloproliferative neoplasms.

Ruxolitinib (INCB018424) is the first JAK1/JAK2 inhibitor approved for treatment of myelofibrosis. JAK/STAT-signalling is known to be involved in the regulation of CD4(+) T cells, which critically orchestrate inflammatory responses. To better understand how ruxolitinib modulates CD4(+) T cell responses, we undertook an in-depth analysis of CD4(+) T cell function upon ruxolitinib exposure. We observed a decrease in total CD3(+) cells after 3 weeks of ruxolitinib treatment in patients with myeloproliferative neoplasms. Moreover, we found that the number of regulatory T cells (Tregs), pro-inflammatory T-helper cell types 1 (Th1) and Th17 were reduced, which were validated by in vitro studies. In line with our in vitro data, we found that inflammatory cytokines [tumour necrosis factor-α (TNF), interleukin (IL)5, IL6, IL1B] were also downregulated in T cells from patients (all P < 0·05). Finally, we showed that ruxolitinib does not interfere with the T cell receptor signalling pathway, but impacts IL2-dependent STAT5 activation. These data provide a rationale for testing JAK inhibitors in diseases triggered by hyperactive CD4(+) T cells, such as autoimmune diseases. In addition, they also provide a potential explanation for the increased infection rates (i.e. viral reactivation and urinary tract infection) seen in ruxolitinib-treated patients.

Impact of allogeneic stem cell transplantation on survival of patients less than 65 years of age with primary myelofibrosis.

Allogeneic hematopoietic stem cell transplantation (SCT) is the only curative option for patients with primary myelofibrosis (PMF), but information on its net advantage over conventional therapies is lacking. Using ad hoc statistical analysis, we determined outcomes in 438 patients <65 years old at diagnosis who received allogenic SCT (n = 190) or conventional therapies (n = 248). Among patients at low risk per the Dynamic International Prognostic Scoring System (DIPSS) model, the relative risk of death after allogenic SCT vs those treated with nontransplant modalities was 5.6 (95% CI, 1.7-19; P = .0051); for intermediate-1 risk it was 1.6 (95% CI, 0.79-3.2; P = .19), for intermediate-2 risk, 0.55 (95% CI, 0.36-0.83; P = .005), and for high risk, 0.37 (95% CI, 0.21-0.66; P = .0007). Thus, patients with intermediate-2 or high-risk PMF clearly benefit from allogenic SCT. Patients at low risk should receive nontransplant therapy, whereas individual counseling is indicated for patients at intermediate-1 risk.

Assessment of bone marrow inflammation in patients with myelofibrosis: an 18F-fluorodeoxyglucose PET/CT study.

PURPOSE: Myelofibrosis is a haematopoietic stem cell neoplasm characterized by bone marrow inflammation, reactive marrow fibrosis and extramedullary haematopoiesis. The aim of this study was to determine if (18)F-FDG PET/CT can be used to noninvasively visualize and quantify the extent and activity of bone marrow involvement. METHODS: In 30 patients, the biodistribution of (18)F-FDG was analysed by measuring the standardized uptake value in the bone marrow compartment and spleen. Imaging findings were compared with laboratory, cytogenetic and histopathological data. RESULTS: Retention of (18)F-FDG was observed in bone marrow and spleen. Bone marrow involvement varied, ranging from mildly increased uptake in the central skeleton to extensive uptake in most parts of the skeleton. The extent of bone marrow involvement decreased over time from initial diagnosis (r s = -0.43, p = 0.019). Metabolic activity of the bone marrow decreased as the histopathological grade of fibrosis increased (r s = -0.37, p = 0.04). There was a significant positive correlation between the metabolic activity of the bone marrow and that of the spleen (p = 0.04). CONCLUSION: (18)F-FDG PET/CT is as a promising technique for the quantitation of bone marrow inflammation in myelofibrosis. Our data indicate that the intensity of bone marrow (18)F-FDG uptake decreases as bone marrow fibrosis increases. Further evaluation in prospective studies is required to determine the potential clinical impact and prognostic significance of PET.

Allogeneic stem cell transplant vs.Janus kinase inhibition in the treatment of primary myelofibrosis or myelofibrosis after essential thrombocythemia or polycythemia vera.

Primary myelofibrosis is one of the Philadelphia chromosome-negative myeloproliferative neoplasms and is the member of that group with the worst survival and the most significant limitations in quality of life. Hepatosplenomegaly due to extramedullary hematopoiesis, constitutional symptoms, and cytopenias are the main manifestations. The natural history is highly variable, and up to 30% of patients can experience acceleration to acute myelogenous leukemia. Conventional therapy is only palliative and not always effective. However, huge advances have been achieved in the past 2 decades toward a better understanding of the pathogenesis of this disease, as well as improved management. Powerful risk stratification systems are now available and can reliably separate the patients into different prognostic categories to aid clinical management. Allogeneic stem cell transplant can offer cure but is still not universally applicable owing to the treatment-related mortality and toxicity. Nevertheless, outcomes of transplant are improving, owing to the introduction of reduced-intensity conditioning regimens and the optimization of remission monitoring techniques and relapse prevention strategies. The discovery of the V617F mutation of JAK2 (Janus kinase 2) and some other molecular aberrations has shed more light on the molecular pathogenesis of the disease and has led to the introduction of novel therapies such as JAK2 inhibitors. In fact, JAK inhibitors have shown promising symptomatic efficacy, and the JAK inhibitor ruxolitinib has also shown a potential survival benefit. Future effort should be made to combine allogeneic stem cell transplant with JAK inhibition.

Dynamic of bone marrow fibrosis regression predicts survival after allogeneic stem cell transplantation for myelofibrosis.

We correlate regression of bone marrow fibrosis (BMF) on day 30 and 100 after dose- reduced allogeneic stem cell transplantation (allo-SCT) in 57 patients with primary or post-essential thrombocythemia/polycythemia vera myelofibrosis with graft function and survival. The distribution of International Prognostic Scoring System (IPSS) risk score categories was 1 patient with low risk, 5 patients with intermediate-1 risk, 18 patients with intermediate-2 risk, and 33 patients with high risk. Before allo-SCT, 41 patients (72%) were classified as XXX [myclofibrosis (MF)]-3 and 16 (28%) were classified as MF-2 according to the World Health Organization criteria. At postengraftment day +30 (±10 days), 21% of the patients had near-complete or complete regression of BMF (MF-0/-1), and on day +100 (±20 days), 54% were MF-0/-1. The 5-year overall survival rate at day +100 was 96% in patients with MF-0/-1 and 57% for those with MF-2/-3 (P = .04). There was no difference in BMF regression at day +100 between IPSS high-risk and low/intermediate-risk patients. Complete donor cell chimerism at day +100 was seen in 81% of patients with MF-0/-1 and in 31% of those with MF-2/-3. Patients with MF-2/-3 at day +100 were more likely to be transfusion-dependent for either RBCs (P = .014) or platelets (P = .018). Rapid BMF regression after reduced-intensity conditioning allo-SCT resulted in a favorable survival independent of IPSS risk score at transplantation.

CD34(+)-selected stem cell boost without further conditioning for poor graft function after allogeneic stem cell transplantation in patients with hematological malignancies.

We retrospectively analyzed outcomes of a CD34(+)-selected stem cell boost (SCB) without prior conditioning in 32 patients (male/22; median age of 54 years; range, 20 to 69) with poor graft function, defined as neutrophils ≤1.5 x 10(9)/L, and/or platelets ≤30 x 10(9)/L, and/or hemoglobin ≤8.5 g/dL). The median interval between stem cell transplantation and SCB was 5 months (range, 2 to 228). The median number of CD34(+) and CD3(+) cells were 3.4 x 10(6)/kg (.96 to 8.30) and 9 x 10(3)/kg body weight (range, 2 to 70), respectively. Hematological improvement was observed in 81% of patients and noted after a median of 30 days (range, 14 to 120) after SCB. The recipients of related grafts responded faster than recipients of unrelated grafts (20 versus 30 days, P = .04). The cumulative incidence of acute (grade II to IV) and chronic graft-versus-host disease (GVHD) after SCB was 17% and 26%, respectively. Patients with acute GVHD received a higher median CD3(+) cell dose. The 2-year probability of overall survival was 45%. We suggest that SCB represents an effective approach to improve poor graft function post transplantation, but optimal timing of SCB administration, anti-infective, and GVHD prophylaxis needs further evaluation.

Serum albumin level predicts survival of patients with gastrointestinal acute graft-versus-host disease after allogeneic stem cell transplantation.

In a retrospective single-centre study, we analysed the prognostic impact of factors identifiable at initial diagnosis of acute GVHD (aGVHD). We retrospectively analysed 495 adult patients of whom 308 (62 %) developed acute GVHD (I-IV) and were included in further analysis. Gut aGVHD was diagnosed in 163/308 cases (53 %). Conditioning was myeloablative conditioning (MAC) in 123 (39.9 %) and reduced intensity (RIC) in 185 (60.1 %) patients. Median serum albumin level at diagnosis of aGVHD was 34 g/l, which was used as cut-off for low vs. normal albumin levels. In patients with gut aGVHD, low albumin level at the time of diagnosis of aGVHD was associated with poorer overall survival (OS) which was 52 vs. 67 % at 1 year and 40 vs. 61 % at 3 years, p = 0.015. In patients with only skin aGVHD, 1- and 3-year OS of patients with low vs. normal albumin levels were 72 vs. 72 % and 59 vs. 57 %, respectively, p = 0.69. In multivariate analysis of patients with gut aGVHD, low serum albumin level ≤34 g/l (relative risk (RR) 2.13, p = 0.003), gut aGVHD grades 3-4 (RR 2.70, p = 0.001), RIC (RR 1.84, p = 0.024), matched unrelated donor (RR 1.86, p = 0.18) and mismatched unrelated donor (RR 2.76, p = 0.03) retained negative impact on OS. Subgroup analysis revealed that impact of albumin was restricted to patients with gut aGVHD after RIC. Low serum albumin levels are associated with poorer OS in patients with gut but not skin aGVHD after RIC but not MAC allogeneic stem cell transplantation.

Allogeneic stem cell transplantation for myelofibrosis with leukemic transformation: a study from the Myeloproliferative Neoplasm Subcommittee of the CMWP of the European Group for Blood and Marrow Transplantation.

Transformed acute myeloid leukemia in myelofibrosis results in a median survival of less than 5 months. We identified 46 of 1048 myelofibrosis patients in the European Group for Blood and Marrow Transplantation registry who received allogeneic stem cell transplantation for acute leukemia evolving from myelofibrosis. The cumulative incidence of treatment-related mortality at 1 year was 28% (95% confidence interval, 14 to 42) and of relapse at 3 years was 47% (95% confidence interval, 31 to 63). The 3-year progression-free (PFS) and overall survival (OS) rates were 26% and 33%, respectively. The only significant factor for survival was complete remission versus no complete remission before transplantation (69% versus 22%, P = .008); however, complete remission was achieved only in 8 patients. Allogeneic stem cell transplantation can cure myelofibrosis patients transformed to leukemia.

Outcome of allogeneic SCT in patients with chronic myeloid leukemia in the era of tyrosine kinase inhibitor therapy.

The introduction of tyrosine kinase inhibitors (TKIs) for chronic myeloid leukemia (CML) led to a dramatic change in the role of allogeneic stem cell transplantation (SCT) with a rapid decline in the number of patients receiving SCT in first chronic phase (CP1). We evaluated 68 consecutive patients in all phases of CML (male/female = 39:29, 27 in CP1), who received SCT from related/unrelated donors (related/unrelated = 23:45) under myeloablative or reduced intensity conditioning (MAC/RIC = 45:23). Forty-eight patients (71 %) received TKIs pre-SCT, 20 patients post-SCT (29 %). Overall survival (OS) of CP1 patients achieved a plateau of 85 % at 10 months. Relapse-free survival (RFS) of CP1 patients was 85 % at 1 and 2 years, and 81 % at 5 years. Multivariate analysis showed adverse OS and RFS for patients transplanted >CP1 (hazard ratio (HR) = 6.61 and 4.62) and those who had grade III-IV aGvHD (HR = 2.45 and 1.82). Patients with advanced CML had estimated OS of 65 and 47 %; and RFS of 41 and 32 % at 1 and 2 years respectively. Therefore, for patients with advanced CML phases, allogeneic SCT provides an acceptable chance of cure. Transplant research should focus on improving conditioning regimens and post-SCT management for this subgroup of CML patients.

Impact of high-risk cytogenetics and achievement of molecular remission on long-term freedom from disease after autologous-allogeneic tandem transplantation in patients with multiple myeloma.

Within a prospective protocol, the incidence and impact of achievement of molecular remission (mCR) and high-risk cytogenetics was investigated in 73 patients with multiple myeloma (MM) after autologous (auto)-allogeneic (allo) tandem stem cell transplantation (SCT). After induction chemotherapy, patients received melphalan 200 mg/m(2) before undergoing auto-SCT, followed 3 months later by melphalan 140 mg/m(2) and fludarabine 180 mg/m(2) before allo-SCT. Sixteen patients had high-risk cytogenetic features, defined by positive FISH for del(17p13) and/or t(4;14). Overall, 66% of the patients achieved CR or near-CR, and 41% achieved mCR, which was sustained negative (at least 4 consecutive samples negative) in 15 patients (21%), with no significant difference in incidence between the patients with high-risk cytogenetics and others (P = .70). After a median follow-up of 6 years, overall 5-year progression-free survival was 29%, with no significant difference between del 17p13/t(4;14)-harboring patients and others (24% versus 30%; P = .70). The 5-year progression-free survival differed substantially according to the achieved remission: 17% for partial remission, 41% for CR, 57% for mCR, and 85% for sustained mCR. These results suggest that auto-allo tandem SCT may overcome the negative prognostic effect of del(17p13) and/or t(4;14) and that achievement of molecular remission resulted in long-term freedom from disease.

Risk models predicting survival after reduced-intensity transplantation for myelofibrosis.

To define a prognostic model for predicting outcome of reduced-intensity allogeneic stem cell transplantation (RIC-ASCT) for myelofibrosis we evaluated 150 homogeneously treated patients and developed a new risk score for overall survival (OS). In a multivariate Cox model for OS, only JAK2 V617F wild-type, age ≥57 years and constitutional symptoms were independently predictive for OS (Hazard Ratio: 2·02; 2·43 and 2·80 respectively). Depending on the presence of one, two or all of these factors, HR of death was 3·08; 4·70 and 16·61 respectively (P < 0·001). This score was compared to the Lille, Cervantes, International Prognostic Scoring System (IPSS), dynamic IPSS (DIPSS) and modified European Blood and Marrow Transplantation Group (EBMT) scores. Lille score correlated significantly with OS but discriminated poorly between the intermediate and high-risk groups (5-year OS 56% and 51% respectively). IPSS and DIPSS correlated significantly with OS but differences between intermediate-1 and intermediate-2 groups were not significant (5-year OS 78% vs. 78% and 70%, 60% respectively). Modified EBMT and Cervantes models did not predict OS post-ASCT. In conclusion, a simple model which includes: age, JAK2 V617F-status and constitutional symptoms can clearly separate distinct risk groups and can be used in addition to the Lille model to predict OS after RIC-ASCT for myelofibrosis.

Chimerism studies with quantitative real-time PCR in stem cell recipients with acute myeloid leukemia.

OBJECTIVE: Chimerism is well-established for surveillance of acute myeloid leukemia (AML) patients after allogeneic hematopoietic stem cell transplantation (HSCT), but interpretation of the results and techniques is not standardized. MATERIALS AND METHODS: We correlated chimerism in 75 AML patients (38 male, 37 female) who underwent myeloablative (n = 36)/reduced (n = 39) allo-HSCT with the risk of relapse and survival. Chimerism was evaluated by quantitative real-time polymerase chain reaction for donor/recipient specific polymorphisms/Y-specific sequences. RESULTS: After HSCT, 40 patients (53%) achieved stable complete donor chimerism (≥ 99.0% of donor alleles), while 35 (47%) failed to achieve stable donor chimerism. Thirty-one patients (41%) showed decreasing donor alleles after having first achieved complete donor chimerism. To investigate the kinetics of mixed chimerism, patients were separated whether they showed subsequent increasing or decreasing donor alleles. Subsequent decrease of donor alleles was associated with relapses in 17 of 18 cases (94%), while no patient with subsequent increasing donor alleles relapsed (p < 0.001). Patients with mixed chimerism and increasing donor alleles had better 2-year disease-free survival (85%) than those with decreasing donor alleles (0%; p < 0.001). CONCLUSIONS: The kinetics of mixed chimerism as assessed by quantitative real-time polymerase chain reaction is an important prognostic predictor in the post-transplantation period of AML patients.

Impact of JAK2V617F mutation status, allele burden, and clearance after allogeneic stem cell transplantation for myelofibrosis.

Allogeneic stem cell transplantation (ASCT) after reduced-intensity conditioning has become a reasonable treatment option for patients with advanced myelofibrosis. The role of characteristic molecular genetic abnormalities, such as JAK2V617F on outcome of ASCT, is not yet elucidated. In 139 of 162 myelofibrosis patients with known JAK2V617F mutation status who received ASCT after reduced-intensity conditioning, the impact of JAK2 genotype, JAK2V617F allele burden, and clearance of mutation after ASCT was evaluated. Overall survival was significantly reduced in multivariate analysis in patients harboring JAK2 wild-type (hazard ratio = 2.14, P = .01) compared with JAK2 mutated patients. No significant influence on outcome was noted for the mutated allele burden analyzed either as continuous variable or after dividing into quartiles. Achievement of JAK2V617F negativity after ASCT was significantly associated with a decreased incidence of relapse (hazard ratio = 0.22, P = .04). In a landmark analysis, patients who cleared JAK2 mutation level in peripheral blood 6 months after ASCT had a significant lower risk of relapse (5% vs 35%, P = .03). We conclude that JAK2V617F-mutated status, but not allele frequency, resulted in an improved survival and rapid clearance after allografting reduces the risk of relapse.

Reduced-intensity conditioning followed by allogeneic hematopoietic stem cell transplantation in myelofibrosis.

Hematopoietic stem cell transplantation offers a curative therapy for patients with myelofibrosis. Because of toxicity, allografting following myeloablative regimens is mainly applicable to young patients. With the introduction of dose-reduced conditioning using busulfan or melphalan with fludarabine, transplantation has become tolerable also for older patients. Implementation of antithymocyte globulin in the conditioning has resulted in effective prevention of graft-versus-host disease and an increased use of alternative donors. Through the discovery of new disease-specific mutations, close monitoring of residual disease became feasible in many patients and the outcome of posttransplant strategies improved. Still challenging is the achievement of new, transplant-derived models to estimate risk status and possible outcome for every individual patient, to help in therapeutic decision making and the determination of the optimal timing of stem cell transplantation. Such a tool may optimally include not only clinicomorphologic characteristics but also other potentially relevant factors such as cytogenetics and novel molecular markers.

Bone marrow mesenchymal stromal cells remain of recipient origin after allogeneic SCT and do not harbor the JAK2V617F mutation in patients with myelofibrosis.

The close association of the myeloproliferative neoplasms with the activating non-receptor tyrosine kinase JAK2V617F mutation is well established. To further clarify the pathomechanisms of this mutation in patients with myelofibrosis, we performed screening with quantitative real-time PCR for the respective mutation in in vitro expanded bone marrow (BM) mesenchymal stromal cells (MSCs) and compared the results with BM/peripheral blood (PB). Eight patients with primary/secondary myelofibrosis were investigated before (n = 4) or after allogeneic stem cell transplantation (n = 4). All patients had systemic evidence of the JAK2V617F mutation in BM/PB (mutation ratios 0.2-23.5) at the time of investigation in contrast to negative results in the MSCs (n = 7) or a very low (0.004) mutation ratio (n = 1) which was probably due to hematopoietic contamination. The four patients post-transplant had systemic donor chimerism between 96.5 and 100% in BM/PB, while MSCs showed no evidence of donor-specific alleles. In conclusion, in myelofibrosis, the JAK2V617F mutation is restricted to hematopoietic cells, and cannot explain the stromal alterations being observed in this disorder. Further, the MSCs remain of recipient origin after allogeneic SCT, which might contribute to the increased risk of graft dysfunction or failure in myelofibrosis patients after allogeneic transplantation.