Chronic inflammation is considered a major driving force for clonal expansion and evolution in the Philadelphia-negative myeloproliferative neoplasms, which include essential thrombocythemia, ...polycythemia vera and primary myelofibrosis (MPNs). One of the key mutation drivers is the JAK2V617F mutation, which has been shown to induce the generation of reactive oxygen species (ROS). Using whole blood gene expression profiling, deregulation of several oxidative stress and anti-oxidative defense genes has been identified in MPNs, including significant downregulation of TP53, the NFE2L2 or NRF2 genes. These genes have a major role for maintaining genomic stability, regulation of the oxidative stress response and in modulating migration or retention of hematopoietic stem cells. Therefore, their deregulation might give rise to increasing genomic instability, increased chronic inflammation and disease progression with egress of hematopoietic stem cells from the bone marrow to seed in the spleen, liver and elsewhere. Interferon-alpha2 (rIFNalpha) is increasingly being recognized as the drug of choice for the treatment of patients with MPNs. Herein, we report the first gene expression profiling study on the impact of rIFNalpha upon oxidative stress and antioxidative defense genes in patients with MPNs (n = 33), showing that rIFNalpha downregulates several upregulated oxidative stress genes and upregulates downregulated antioxidative defense genes. Treatment with rIFNalpha induced upregulation of 19 genes in ET and 29 genes in PV including CXCR4 and TP53. In conclusion, this rIFNalpha- mediated dampening of genotoxic damage to hematopoietic cells may ultimately diminish the risk of additional mutations and accordingly clonal evolution and disease progression towards myelofibrotic and leukemic transformation.
We report the final 2-year end-of-study results from the first clinical trial investigating combination treatment with ruxolitinib and low-dose pegylated interferon-α2 (PEG-IFNα2). The study included ...32 patients with polycythemia vera and 18 with primary or secondary myelofibrosis; 46 patients were previously intolerant of or refractory to PEGIFNα2. The primary outcome was efficacy, based on hematologic parameters, quality of life measurements, and JAK2 V617F allele burden. We used the 2013 European LeukemiaNet and International Working Group- Myeloproliferative Neoplasms Research and Treatment response criteria, including response in symptoms, splenomegaly, peripheral blood counts, and bone marrow. Of 32 patients with polycythemia vera, ten (31%) achieved a remission which was a complete remission in three (9%) cases. Of 18 patients with myelofibrosis, eight (44%) achieved a remission; five (28%) were complete remissions. The cumulative incidence of peripheral blood count remission was 0.85 and 0.75 for patients with polycythemia vera and myelofibrosis, respectively. The Myeloproliferative Neoplasm Symptom Assessment Form total symptom score decreased from 22 95% confidence interval (95% CI):, 16-29 at baseline to 15 (95% CI: 10-22) after 2 years. The median JAK2 V617F allele burden decreased from 47% (95% CI: 33-61%) to 12% (95% CI: 6-22%), and 41% of patients achieved a molecular response. The drop-out rate was 6% among patients with polycythemia vera and 32% among those with myelofibrosis. Of 36 patients previously intolerant of PEG-IFNα2, 31 (86%) completed the study, and 24 (67%) of these received PEG-IFNα2 throughout the study. In conclusion, combination treatment improved cell counts, reduced bone marrow cellularity and fibrosis, decreased JAK2 V617F burden, and reduced symptom burden with acceptable toxicity in several patients with polycythemia vera or myelofibrosis. #EudraCT2013-003295-12.
Introduction The Philadelphia chromosome-negative myeloproliferative neoplasms are a group of slowly progressing haematological malignancies primarily characterised by an overproduction of myeloid ...blood cells. Patients are treated with various drugs, including the JAK1/2 inhibitor ruxolitinib. Mathematical modelling can help propose and test hypotheses of how the treatment works. Materials and methods We present an extension of the Cancitis model, which describes the development of myeloproliferative neoplasms and their interactions with inflammation, that explicitly models progenitor cells and can account for treatment with ruxolitinib through effects on the malignant stem cell response to cytokine signalling and the death rate of malignant progenitor cells. The model has been fitted to individual patients’ data for the JAK2 V617F variant allele frequency from the COMFORT-II and RESPONSE studies for patients who had substantial reductions (20 percentage points or 90% of the baseline value) in their JAK2 V617F variant allele frequency ( n = 24 in total). Results The model fits very well to the patient data with an average root mean square error of 0.0249 (2.49%) when allowing ruxolitinib treatment to affect both malignant stem and progenitor cells. This average root mean square error is much lower than if allowing ruxolitinib treatment to affect only malignant stem or only malignant progenitor cells (average root mean square errors of 0.138 (13.8%) and 0.0874 (8.74%), respectively). Discussion Systematic simulation studies and fitting of the model to the patient data suggest that an initial reduction of the malignant cell burden followed by a monotonic increase can be recapitulated by the model assuming that ruxolitinib affects only the death rate of malignant progenitor cells. For patients exhibiting a long-term reduction of the malignant cells, the model predicts that ruxolitinib also affects stem cell parameters, such as the malignant stem cells’ response to cytokine signalling.
The neutrophil-to-lymphocyte ratio(NLR) is increased in chronic inflammation and myeloproliferative neoplasms (MPN). We hypothesize that NLR is associated with all-cause mortality and mortality by ...comorbidity burden in the general population and individuals with MPN. We included 835,430 individuals from The Danish General Suburban Population Study, general practitioners, and outpatient clinics. We investigated NLR on mortality stratified by prevalent and incident MPN, essential thrombocythemia (ET), polycythemia vera (PV), myelofibrosis (MF), comorbidity burden (CCI-score), and the Triple-A risk score using hazard ratio (HR) and 95% confidence interval (95%CI). NLR 1-1.9 was the reference level. During a median follow-up of 11.2 years, 197,802 deaths were recorded. All-cause mortality increased for a stepwise increasing NLR with a HR (95%CI) for NLR ≥ 6 of 2.06(2.03-2.09) for the whole population and 2.93(2.44-3.50) in prevalent MPN. ET, PV, and MF had a HR (95%CI) for NLR ≥ 2 of 2.14(1.71-2.69), 2.19(1.89-2.54), and 2.31(1.91-2.80). Results were similar for incident MPN. Mortality was higher for stepwise increasing NLR and CCI-score(p
< 2×10
), with a HR for NLR ≥ 6 of 2.23(2.17-2.29), 4.10(4.01-4.20), and 7.69(7.50-7.89), for CCI-score 0, 1-2, or ≥3. The Triple-A risk score demonstrated alignment with NLR. Increasing NLR and comorbidity burden were associated with lower survival in individuals without MPN but were even worse in prevalent and incident MPN, ET, PV, and MF.
ABSTRACT
Essential thrombocythemia (ET) is part of the Philadelphia chromosome-negative myeloproliferative neoplasms. It is characterized by an increased risk of thromboembolic events and also to a ...certain degree hypermetabolic symptoms. The gut microbiota is an important initiator of hematopoiesis and regulation of the immune system, but in patients with ET, where inflammation is a hallmark of the disease, it is vastly unexplored. In this study, we compared the gut microbiota via amplicon-based 16S rRNA gene sequencing of the V3–V4 region in 54 patients with ET according to mutation status Janus-kinase 2 (
JAK2V617F
)-positive vs
JAK2V617F
-negative patients with ET, and in 42 healthy controls (HCs). Gut microbiota richness was higher in patients with ET (median-observed richness, 283.5; range, 75–535) compared with HCs (median-observed richness, 191.5; range, 111–300;
P
< 0.001). Patients with ET had a different overall bacterial composition (beta diversity) than HCs (analysis of similarities ANOSIM;
R
= 0.063,
P
= 0.004). Patients with ET had a significantly lower relative abundance of taxa within the
Firmicutes
phylum compared with HCs (51% vs 59%,
P
= 0.03), and within that phylum, patients with ET also had a lower relative abundance of the genus
Faecalibacterium
(8% vs 15%,
P
< 0.001), an important immunoregulative bacterium. The microbiota signatures were more pronounced in patients harboring the
JAK2V617F
mutation, and highly similar to patients with polycythemia vera as previously described. These findings suggest that patients with ET may have an altered immune regulation; however, whether this dysregulation is induced in part by, or is itself inducing, an altered gut microbiota remains to be investigated.
IMPORTANCE
Essential thrombocythemia (ET) is a cancer characterized by thrombocyte overproduction. Inflammation has been shown to be vital in both the initiation and progression of other myeloproliferative neoplasms, and it is well known that the gut microbiota is important in the regulation of our immune system. However, the gut microbiota of patients with ET remains uninvestigated. In this study, we characterized the gut microbiota of patients with ET compared with healthy controls and thereby provide new insights into the field. We show that the gut microbiota of patients with ET differs significantly from that of healthy controls and the patients with ET have a lower relative abundance of important immunoregulative bacteria. Furthermore, we demonstrate that patients with
JAK2V617F
-positive ET have pronounced gut microbiota signatures compared with
JAK2V617F
-negative patients. Thereby confirming the importance of the underlying mutation, the immune response as well as the composition of the microbiota.
Essential thrombocythemia (ET) is a cancer characterized by thrombocyte overproduction. Inflammation has been shown to be vital in both the initiation and progression of other myeloproliferative neoplasms, and it is well known that the gut microbiota is important in the regulation of our immune system. However, the gut microbiota of patients with ET remains uninvestigated. In this study, we characterized the gut microbiota of patients with ET compared with healthy controls and thereby provide new insights into the field. We show that the gut microbiota of patients with ET differs significantly from that of healthy controls and the patients with ET have a lower relative abundance of important immunoregulative bacteria. Furthermore, we demonstrate that patients with
JAK2V617F
-positive ET have pronounced gut microbiota signatures compared with
JAK2V617F
-negative patients. Thereby confirming the importance of the underlying mutation, the immune response as well as the composition of the microbiota.
BackgroundThe majority of JAK2V617F-negative myeloproliferative neoplasms (MPN) have disease-initiating frameshift mutations in calreticulin (CALR) resulting in a common novel C-terminal mutant ...fragment (CALRMUT), representing an attractive source of neoantigens for cancer vaccines. However, studies have shown that CALRMUT-specific T cells are rare in CALRMUT MPN patients, but the underlying reasons for this phenomenon are unknown.MethodsIn this study, we examine class-I major histocompatibility complex (MHC-I) allele frequency in CALRMUT MPN patients from two independent cohorts and observed that MHC-I alleles that present CALRMUT neoepitopes with high affinity are under-represented in CALRMUT MPN patients. We speculate that this is due to an increased chance of immune-mediated tumor rejection by individuals expressing one of these MHC-I alleles such that the disease never clinically manifests. As a consequence of this MHC-I allele restriction, we reasoned that CALRMUT MPN patients would not efficiently respond to cancer vaccines composed of the CALRMUT fragment, but could do so when immunized with a properly modified CALRMUT heteroclitic peptide vaccine approach.ResultsWe found that heteroclitic CALRMUT peptides specifically designed for CALRMUT MPN patient MHC-I alleles efficiently elicited a cross-reactive CD8+ T cell response in human PBMC samples otherwise unable to respond to the matched weakly immunogenic CALRMUT native peptides. We also modeled this effect in mice and observed that C57BL/6J mice, which are unable to mount an immune response to the human CALRMUT fragment, can mount a cross-reactive CD8+ T cell response against a CALRMUT-derived peptide upon heteroclitic peptide immunization and this was further amplified by combining the heteroclitic peptide vaccine with blockade of the immune checkpoint molecule PD-1.ConclusionsTogether, our data underscore the therapeutic potential of heteroclitic peptide-based cancer vaccines in CALRMUT MPN patients.Ethics ApprovalApproval was obtained for the use of patient-derived specimens and access to clinical data extracted from patient charts by the Institutional Review Boards at Memorial Sloan Kettering Cancer Center, the Dana-Farber Cancer Institute and the Massachusetts General Hospital, as well as by the Danish Regional Science Ethics Committee. Mouse experiments were performed in accordance with institutional guidelines under a protocol approved by the Memorial Sloan-Kettering Cancer Center Institutional Animal Care and Use Committee.
Neutrophil extracellular traps (NETs) may play a pathogenic role in the thrombosis associated with myeloproliferative neoplasms (MPNs). We measured serum NET levels in 128 pretreatment samples from ...patients with MPNs and in 85 samples taken after 12 months of treatment with interferon alpha‐2 (PEG‐IFNα‐2) formulations or hydroxyurea (HU). No differences in NET levels were observed across subdiagnoses or phenotypic driver mutations. In PV, a JAK2V617F+ allele burden ≥50% associated with increased NET levels (p = 0.006). Baseline NET levels correlated with neutrophil count (r = 0.29, p = 0.001), neutrophil‐to‐lymphocyte ratio (r = 0.26, p = 0.004) and JAK2V617F allele burden (r = 0.22, p = 0.03), particularly in patients with PV and with allele burden ≥50% (r = 0.50, p = 0.01, r = 0.56, p = 0.002 and r = 0.45, p = 0.03 respectively). In PV, after 12 months of treatment, NET levels decreased on average by 60% in patients with allele burden ≥50%, compared to only 36% in patients with an allele burden <50%. Overall, treatment with PEG‐IFNα‐2a or PEG‐IFNα‐2b reduced NETs levels in 77% and 73% of patients, respectively, versus only 53% of HU‐treated patients (average decrease across treatments: 48%). Normalization of blood counts did not per se account for these reductions. In conclusion, baseline NET levels correlated with neutrophil count, NLR and JAK2V617F allele burden, and IFNα was more effective at reducing prothrombotic NET levels than HU.
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