Efficacy and safety outcomes in Japanese patients with low-risk polycythemia vera treated with ropeginterferon alfa-2b.

Polycythemia vera (PV) is a Philadelphia chromosome-negative myeloproliferative neoplasm characterized by clonal erythrocytosis. A phase 2 study reported that ropeginterferon alfa-2b is a well-tolerated and effective treatment for PV in Japanese patients. This post hoc analysis of the phase 2 data further evaluated outcomes in patients at low risk of thrombosis (low-risk PV). Among 20 patients with low-risk PV, 60.0% (12/20) and 85.0% (17/20) achieved < 45% hematocrit by weeks 24 and 52, respectively. The proportion of responders with complete hematologic response (CHR) was 60.0% (12/20) at week 52, and the median time to response was 11.9 months. The mean JAK2 V617F allele burden decreased from 75.8% at baseline to 53.7% at week 52. No patient experienced thrombosis or bleeding episodes. All patients experienced treatment-emergent adverse events (TEAEs) related to ropeginterferon alfa-2b, but no grade ≥ 3 TEAEs or deaths related to ropeginterferon alfa-2b occurred, and no new safety concerns arose. This analysis indicated that ropeginterferon alfa-2b may be an effective treatment option for Japanese patients with low-risk PV.

[Recent findings and advances in treatment of myeloproliferative neoplasms].

Many novel agents have been developed for BCR::ABL1-negaive myeloproliferative neoplasms (MPN), namely, polycythemia vera (PV), essential thrombocythemia (ET), and primary myelofibrosis (PMF). Some of these agents not only achieve hematologic complete response, reduce spleen size, and alleviate constitutional symptoms, but also induce molecular response, which means that they reduce the allele burden of driver gene mutations. These agents also prevent and alleviate fibrosis in bone marrow, which reduces the incidence of thrombotic events and disease progression and might improve prognosis. This article discusses the latest findings and promising treatments, including ongoing clinical trials, in PV, ET, and PMF.

Effective Management of Polycythemia Vera With Ropeginterferon Alfa-2b Treatment.

Background: Polycythemia vera (PV) is a myeloproliferative neoplasm. Ropeginterferon alfa-2b is a new-generation polyethylene glycol-conjugated proline-interferon. It is approved for the treatment of PV at a starting dose of 100 µg (50 µg for patients receiving hydroxyurea (HU)) and dose titrations up to 500 µg by 50 µg increments. The study was aimed at assessing its efficacy and safety at a higher starting dose and simpler intra-patient dose escalation. Methods: Forty-nine patients with PV having HU intolerance from major hospitals in China were treated biweekly with an initial dose of 250 µg, followed by 350 µg and 500 µg thereafter if tolerated. Complete hematological response (CHR) was assessed every 12 weeks based on the European LeukemiaNet criteria. The primary endpoint was the CHR rate at week 24. The secondary endpoints included CHR rates at weeks 12, 36 and 52, changes of JAK2V617F allelic burden, time to first CHR, and safety assessments. Results: The CHR rates were 61.2%, 69.4% and 71.4% at weeks 24, 36, and 52, respectively. Mean allele burden of the driver mutation JAK2V617F declined from 58.5% at baseline to 30.1% at 52 weeks. Both CHR and JAK2V617F allele burden reduction showed consistent increases over the 52 weeks of the treatment. Twenty-nine patients (63.0%) achieved partial molecular response (PMR) and two achieved complete molecular response (CMR). The time to CHR was rapid and median time was 5.6 months according to central lab results. The CHRs were durable and median CHR duration time was not reached at week 52. Mean spleen index reduced from 55.6 cm2 at baseline to 50.2 cm2 at week 52. Adverse events (AEs) were mostly mild or moderate. Most common AEs were reversible alanine aminotransferase and aspartate aminotransferase increases, which were not associated with significant elevations in bilirubin levels or jaundice. There were no grade 4 or 5 AEs. Grade 3 AEs were reversible and manageable. Only one AE led to discontinuation. No incidence of thromboembolic events was observed. Conclusion: The 250-350-500 µg dosing regimen was well tolerated and effectively induced CHR and MR and managed spleen size increase. Our findings demonstrate that ropeginterferon alfa-2b at this dosing regimen can provide an effective management of PV and support using this dosing regimen as a treatment option.

Association of JAK2V617F allele burden and clinical correlates in polycythemia vera: a systematic review and meta-analysis.

Janus kinase 2 (JAK2) V617F mutation is present in most patients with polycythemia vera (PV). One persistently puzzling aspect unresolved is the association between JAK2V617F allele burden (also known as variant allele frequency) and the relevant clinical characteristics. Numerous studies have reported associations between allele burden and both hematologic and clinical features. While there are strong indications linking high allele burden in PV patients with symptoms and clinical characteristics, not all associations are definitive, and disparate and contradictory findings have been reported. Hence, this study aimed to synthesize existing data from the literature to better understand the association between JAK2V617F allele burden and relevant clinical correlates. Out of the 1,851 studies identified, 39 studies provided evidence related to the association between JAK2V617F allele burden and clinical correlates, and 21 studies were included in meta-analyses. Meta-analyses of correlation demonstrated that leucocyte and erythrocyte counts were significantly and positively correlated with JAK2V617F allele burden, whereas platelet count was not. Meta-analyses of standardized mean difference demonstrated that leucocyte and hematocrit were significantly higher in patients with higher JAK2V617F allele burden, whereas platelet count was significantly lower. Meta-analyses of odds ratio demonstrated that patients who had higher JAK2V617F allele burden had a significantly greater odds ratio for developing pruritus, splenomegaly, thrombosis, myelofibrosis, and acute myeloid leukemia. Our study integrates data from approximately 5,462 patients, contributing insights into the association between JAK2V617F allele burden and various hematological parameters, symptomatic manifestations, and complications. However, varied methods of data presentation and statistical analyses prevented the execution of high-quality meta-analyses.

Exposure-efficacy and exposure-safety analyses of ropeginterferon alfa-2b treatment in patients with polycythaemia vera.

AIMS: To investigate the exposure-response (E-R) relationship, including exposure-efficacy and exposure-safety, of ropeginterferon alfa-2b treatment in patients with polycythaemia vera (PV). METHODS: Based on the results of the phase II trial A20-202 regarding ropeginterferon alfa-2b in patients with PV, E-R analyses were performed to evaluate the efficacy and safety of the given dosing regimen. The E-R analyses were based on logistic and linear regression and the relationship between exposure to ropeginterferon alfa-2b and key efficacy and safety variables. The key efficacy variables included complete haematologic response (CHR) and reduction of the driver mutation JAK2V617F. The safety variable was treatment-related adverse events (TRAEs). RESULTS: A clear relationship between the exposure to ropeginterferon alfa-2b and CHR was observed, with an increase in drug exposure resulting in an increased probability of achieving CHR. Similar CHR probabilities were observed in the third and fourth quantiles of the average concentration at Week 24. The results from the exposure-JAK2V617F model indicated that the JAK2V617F allele burden decreased with increasing exposure to ropeginterferon alfa-2b and baseline body surface area. Exposure-safety analysis revealed a risk of AEs associated with transaminase abnormalities, which were not associated with clinical significance. CONCLUSIONS: Our analyses have shown that patients with PV treated with ropeginterferon alfa-2b had an increased probability of achieving CHR and a molecular response with acceptable safety risks at the 250-350-500 µg titration dosing regimen. This study has provided the relevant data for the application of a biologics licence of ropeginterferon alfa-2b for PV treatment in China.

Recurrent Stroke with Rapid Development of Intracranial Artery Stenosis and Subsequent Successful Mechanical Thrombectomy in Essential Thrombocythemia.

Essential thrombocythemia is a myeloproliferative neoplasm. Ischemic stroke is frequently the first manifestation of essential thrombocythemia. We herein report a patient with JAK2V617 mutation-positive essential thrombocythemia who developed recurrent ischemic stroke with rapid development of intracranial artery stenosis and subsequently underwent successful mechanical thrombectomy. The high JAK2V617F allele burden in our patient (58.4%) may have affected the patient's condition. We discuss similar reports in the literature and the possible pathophysiologic mechanism of large artery involvement in these patients.

Clinical Impact of JAK2V617F Allele Burden in Philadelphia-Negative Myeloproliferative Neoplasms

Objective: The impact of JAK2V617F allele burden on clinical course in Philadelphia-negative (Ph-negative) myeloproliferative neoplasms (MPNs) is not clear. We analyzed the clinical impact of JAK2V617F allele burden in a relatively large series of patients with Ph-negative MPNs and long-term follow-up. Materials and Methods: A total of 228 patients with Ph-negative MPNs, including 118 with essential thrombocythemia (ET), 84 with primary myelofibrosis (PMF), and 26 with polycythemia vera (PV), were analyzed. The JAK2 MutaScreen assay was used to quantify JAK2V617F allele burden in genomic DNA. Results: In PV cases, high JAK2V617F allele burden was associated with a trend towards inferior overall survival. In ET, high JAK2V617F allele burden was associated with lower hemoglobin and hematocrit levels, higher lactate dehydrogenase (LDH) levels, larger spleen size, and increased bleeding and mortality rates. In PMF, high JAK2V617F allele burden was associated with higher leukocyte counts and larger spleen size. In the entire cohort, high allele burden was associated with higher leukocyte and lower platelet counts, higher LDH levels, larger spleen size, higher percentage of bleeding events, higher death rate, and inferior overall survival. Conclusion: Our results suggest that high JAK2V617F allele burdens are associated with more severe disease in PV and ET. In PMF, high JAK2V617F allele burdens were associated with more pronounced myeloproliferative phenotypes. In Ph-negative MPNs, high allele burdens were associated with more aggressive phenotypes. Our data with a long follow-up period support the possibility of JAK2V617F allele burden being used as a marker for predicting clinical phenotype in cases of Ph-negative MPNs.

A new dosing regimen of ropeginterferon alfa-2b is highly effective and tolerable: findings from a phase 2 study in Chinese patients with polycythemia vera.

Ropeginterferon alfa-2b represents a new-generation pegylated interferon-based therapy and is administered every 2-4 weeks. It is approved for polycythemia vera (PV) treatment in the United States and Europe with a starting dose of 100 µg (50 µg for patients receiving hydoxyurea) and intra-patient dose titrations up to 500 µg at 50 µg increments, which took approximately 20 or more weeks to reach a plateau dose level. This study aimed to assess ropeginterferon alfa-2b at an alternative dosing regimen with a higher starting dose and quicker intra-patient dose titrations, i.e., the 250-350-500 µg schema, in 49 Chinese patients with PV with resistance or intolerance to hydroxyurea. The primary endpoint of the complete hematologic response rate at treatment weak 24 was 61.2%, which was notably higher than 43.1% at 12 months with the approved dosing schema. The JAK2V617F allele burden decreased from baseline to week 24 (17.8% ± 18.0%), with one patient achieving a complete molecular response. Ropeginterferon alfa-2b was well-tolerated and most adverse events (AEs) were mild or moderate. Common AEs included alanine aminotransferase and aspartate aminotransferase increases mostly at grade 1 or 2 levels. Patients did not present with jaundice or significant bilirubin level increase. No grade 4 or 5 AEs occurred. Seven patients (14.3%) experienced reversible, drug-related grade 3 AEs. No AEs led to treatment discontinuation. Ropeginterferon alfa-2b at the 250-350-500 µg regimen is highly effective and well-tolerated and can help patients achieve greater and rapid complete hematologic and molecular responses.Clinical Trial Registration: This trial is registered at ClinicalTrials.gov (Identifier: NCT05485948) and in China (China National Medical Products Administration Registration Number: CTR20211664).

Allelic burden of Janus kinase 2 in a 6-month course of therapy for myeloproliferative neoplasms.

BACKGROUND: Janus kinase 2 (JAK2) V617F gene mutation is an important marker for the diagnosis of Philadelphia negative Myeloproliferative neoplasms (MPN) which is subdivided into Polycythemia Vera (PV), Primary Myelofibrosis (PMF), and Essential Thrombocythemia (ET). The aim here is to investigate the JAK2 allele burden of the patients diagnosed with the subgroups of MPN and to demonstrate the alterations of hematological parameters and spleen size between diagnosis and 6 months of treatment. METHODS: A total of 107 patients with the diagnosis of MPN and negative Philadelphia chromosome, 51 males and 56 females with a mean age of 59,74 ± 16,41 years, were included in the study. Diagnosis of MPN was based on the World Health Organization (WHO) criteria. Subgroups of MPN distributed as 49,5% ET, 46,7% PV, and 3,8% PMF. Findings such as the age of the patients, JAK-2 allele burden, and laboratory findings of splenomegaly were examined at the time of diagnosis, 3rd month, and 6th month. JAK2 allele burden and spleen size were re-evaluated in 6th month. RESULTS: Our study confirmed the findings of high Hb, HCT, and RBC but low platelet values in PV patients with high JAK2 allele burden with respect to other groups, a positive correlation between JAK2 allele burden and LDH. CONCLUSIONS: A novel finding of our study is, that there is not any reducing effect of the phlebotomy on JAK2 allele burden in PV patients whether they receive phlebotomy or not. Evaluation of the spleen size alteration during 6 months within the subgroups demonstrated a decrease in PV and ET groups whereas no statistically significant difference was found in the PMF group.

Comparison of Clinical and Molecular Features Between Patients With Essential Thrombocythemia and Early/Prefibrotic Primary Myelofibrosis Presenting With Thrombocytosis in Taiwan.

OBJECTIVES: The clinical presentations of essential thrombocythemia (ET) may be quite similar to early/prefibrotic primary myelofibrosis (pre-PMF), especially in pre-PMF presenting with thrombocytosis (pre-PMF-T), but may be associated with a different outcome. It is very important to distinguish these two entities. The aim of this study was to address the clinical and prognostic relevance of distinguishing pre-PMF-T from ET. METHODS: All patients, including 258 with ET and 105 with pre-PMF-T, received JAK2V617F, MPL (exon 10), and CALR (exon 9) mutation analysis and allele burden measurement for JAK2V617F and CALR mutants. RESULTS: Patients with pre-PMF-T had an older age and higher leukocyte and platelet counts but lower hemoglobin levels than patients with ET. Patients with pre-PMF-T had a shorter overall, leukemia-free, and thrombosis-free survival compared with patients with ET. Patients with ET had a higher rate of cerebral ischemic stroke, whereas patients with pre-PMF-T tended to have splanchnic vein thrombosis. The frequencies of JAK2V617F, CALR, and MPL mutations and CALR allele burden were no different, but JAK2V617F allele burden was significantly higher in pre-PMF-T. Patients with pre-PMF-T with the JAK2V617F mutation had an inferior overall survival and thrombosis-free survival, whereas the status of driver gene mutations did not influence the outcomes of patients with ET. CONCLUSIONS: ET and pre-PMF-T were two distinct disease entities and exhibited different clinical phenotype, genotype, and outcomes.

Essential thrombocythaemia progression to the fibrotic phase is associated with a decrease in JAK2 and PDL1 levels.

It has been postulated that the changes in the molecular characteristics of the malignant clone(s) and the abnormal activation of JAK-STAT signaling are responsible for myeloproliferative neoplasm progression to more advanced disease phases and the immune escape of the malignant clone. The continuous JAK-STAT pathway activation leads to enhanced activity of the promoter of CD274 coding programmed death-1 receptor ligand (PD-L1), increased PD-L1 level, and the immune escape of MPN cells. The aim of study was to evaluate the PDL1 mRNA and JAK2 mRNA level in molecularly defined essential thrombocythaemia (ET) patients (pts) during disease progression to post-ET- myelofibrosis (post-ET-MF). The study group consisted of 162 ET pts, including 30 pts diagnosed with post-ET-MF. The JAK2V617F, CALR, and MPL mutations were found in 59.3%, 19.1%, and 1.2% of pts, respectively. No copy-number alternations of the JAK2, PDL1, and PDCDL1G2 (PDL2) genes were found. The level of PD-L1 was significantly higher in the JAK2V617F than in the JAK2WT, CALR mutation-positive, and triple-negative pts. The PD-L1 mRNA level was weakly correlated with both the JAK2V617F variant allele frequency (VAF), and with the JAK2V617F allele mRNA level. The total JAK2 level in post-ET-MF pts was lower than in ET pts, despite the lack of differences in the JAK2V617F VAF. In addition, the PD-L1 level was lower in post-ET-MF. A detailed analysis has shown that the decrease in JAK2 and PDL1 mRNA levels depended on the bone marrow fibrosis grade. The PDL1 expression showed no differences in relation to the genotype of the JAK2 haplotypeGGCC_46/1, hemoglobin concentration, hematocrit value, leukocyte, and platelet counts. The observed drop of the total JAK2 and PDL1 levels during the ET progression to the post-ET-MF may reflect the changes in the JAK2V617F positive clone proliferative potential and the PD-L1 level-related immunosuppressive effect. The above-mentioned hypothesis is supported by The Cancer Genome Atlas (TCGA) data, confirming a strong positive association between CD274 (encoding PD-L1), CXCR3 (encoding CXCR3), and CSF1 (encoding M-CSF) expression levels, and recently published results documenting a drop in the CXCR3 level and circulating M-CSF in patients with post-ET-MF.

von Willebrand factor activity levels are influenced by driver mutation status in polycythemia vera and essential thrombocythemia patients with well-controlled platelet counts.

von Willebrand factor ristocetin cofactor (vWF activity) and platelet count (PLT) are negatively correlated in patients with polycythemia vera (PV) and essential thrombocythemia (ET). However, vWF activity does not always normalize upon controlling PLT in those patients. To address this issue, we investigated the correlation between vWF activity and PLT in PV and ET patients. The negative correlation between vWF activity and PLT was stronger in calreticulin mutation-positive (CALR+) ET than in Janus kinase 2 mutation-positive (JAK2+) PV or ET groups. When PLT were maintained at a certain level (<600 × 109 /L), low vWF activity (<50%) was more frequently observed in JAK2+ PV patients than in JAK2+ ET (p = .013) or CALR+ ET (p = .013) groups, and in PV and ET patients with ≥50% JAK2+ allele burden than in those with allele burden <50% (p = .015). High vWF activity (>150%) was more frequent in the JAK2+ ET group than in the CALR+ ET group (p = .005), and often associated with vasomotor symptoms (p = .002). This study suggests that some patients with JAK2+ PV or ET have vWF activity outside the standard range even with well-controlled PLT, and that the measurement of vWF activity is useful for assessing the risk of thrombosis and hemorrhage.

Impact of Anti-Endothelial Cell Antibodies (AECAs) in Patients with Polycythemia Vera and Thrombosis.

Polycythemia vera (PV) causes thrombosis. Erythrocytosis and cell adhesiveness are responsible for thrombosis. JAK2V617F causes inflammation and autoimmunity; however, whether or not autoimmunity or inflammation causes thrombosis has yet to be proven. In 60 PV patients, we analyzed JAK2V671F and its allele burden, autoimmune Th17 cells, interleukin-17 (IL-17), anti-endothelial cell antibodies (AECAs), endothelial leukocyte adhesion molecule-1 (ELAM-1), intercellular adhesion molecule-1 (ICAM-1), and von Willebrand factor antigen (VWF: Ag). Fifty blood donors were used as the controls. All patients were on phlebotomy-maintaining hematocrit <45% and aspirin. Of the 60 patients, 40 had thrombosis. Those patients with thrombosis had a higher JAK2V617F allele burden than those without thrombosis, andTh17 cells and IL-17 were also higher in patients with thrombosis. Interestingly, we observed a high AECA IgG ELISA ratio (ER) in patients with thrombosis, which was normal in patients without thrombosis. We found high ELAM-1 and ICAM-1 as well as high VWF:Ag in patients with thrombosis compared to patients without thrombosis. AECA-positive sera from patients with thrombosis showed enhanced binding to cytokine-treated HUVEC and a positive antibody-dependent cellular cytotoxicity, suggesting that AECA may contribute to vascular injury. A positive correlation between AECAs, allele burden, and thrombosis was found. These results suggest that autoimmunity may be an additional mechanism in PV thrombogenesis.

Efficacy and safety of ropeginterferon alfa-2b in Japanese patients with polycythemia vera: an open-label, single-arm, phase 2 study.

Ropeginterferon alfa-2b is a novel, site-selective, monopegylated recombinant human interferon alfa-2b. Safety and efficacy of ropeginterferon alfa-2b for the treatment of polycythemia vera were demonstrated in clinical studies conducted in European countries, but clinical studies in Japanese patients are lacking. This phase 2, open-label, multicenter, single-arm study investigated the safety and efficacy of ropeginterferon alfa-2b in 29 Japanese patients with polycythemia vera including young patients and patients with low thrombosis risk who are difficult to receive guideline-based standard treatments. The primary outcome of durable complete hematologic response without phlebotomy at months 9 and 12 was achieved by 8/29 (27.6%) patients. The fastest complete hematologic response was observed at week 12. A corresponding reduction in the JAK2 V617F allele burden from baseline to 52 weeks was also observed (mean ± standard deviation = - 19.2% ± 22.6%). No new safety concerns were identified in Japanese patients when compared with previous studies of ropeginterferon alfa-2b in European populations; the most common treatment-related adverse events were alopecia (55.2%), fatigue (27.6%) and influenza-like illness (27.6%). Most treatment-related adverse events were mild or moderate, with none of grade ≥ 3. Ropeginterferon alfa-2b is a safe and efficacious treatment option in Japanese patients with polycythemia vera.

Standards of Genetic Testing in the Diagnosis and Prognostication of Systemic Mastocytosis in 2022: Recommendations of the EU-US Cooperative Group.

Mastocytosis comprises rare heterogeneous diseases characterized by an increased accumulation of abnormal mast cells in various organs/tissues. The pathogenesis of mastocytosis is strongly linked to the presence of KIT-activating mutations. In systemic mastocytosis (SM), the most frequent mutation encountered is KIT p.D816V, whose presence constitutes one of the minor diagnostic criteria. Different techniques are used to search and quantify the KIT p.D816V mutant; however, allele-specific quantitative PCR and droplet digital PCR are today the most sensitive. The analysis of the KIT p.D816V allele burden has undeniable interest for diagnostic, prognostic, and therapeutic monitoring. The analysis of non-mast cell hematological compartments in SM is similarly important because KIT p.D816V multilineage involvement is associated with a worse prognosis. In addition, in advanced forms of SM, mutations in genes other than KIT are frequently identified and affect negatively disease outcome and response to therapy. Thus, combined quantitative and sensitive analysis of KIT mutations and next-generation sequencing of other recurrently involved myeloid genes make it possible to better characterize the extent of the affected cellular compartments and additional molecular aberrations, providing a more detailed overview of the complex mutational landscape of SM, in relation with the clinical heterogeneity of the disease. In this article, we report the latest recommendations of the EU-US Cooperative Group presented in September 2020 in Vienna during an international working conference, on the techniques we consider standard to detect and quantify the KIT p.D816V mutant in SM and additional myeloid mutations found in SM subtypes.

[Evaluation of the correlation between JAK2V617F allele burden by Quenching Probe-Tm method and allele-specific quantitative PCR method].

The JAK2V617F mutation is a driver mutation of myeloproliferative neoplasms (MPNs). V617F allele burden is considered a risk factor for complications associated with MPNs and is a predictor of prognosis. In Japan, V617F allele burden has been measured in laboratory settings using the i-densyTM IS-5320 genetic analyzer with the quenching probe-Tm (QP-Tm) method. However, since 2020, allele-specific quantitative PCR (AS-qPCR) is being performed in clinical settings for measuring V617F allele burden. To investigate the clinical usefulness of the QP-Tm method in patients with MPNs, we evaluated the V617F allele burden measured by both the methods. A good correlation was observed between the V617F allele burden determined using QP-Tm and that determined using AS-qPCR (P<0.001, rs=0.952). The median mutant allele burden, as determined using the QP-Tm method, was significantly higher in patients with polycythemia vera than in those with essential thrombocythemia. The results of this study suggested that the QP-Tm method will continue to be useful clinical ancillary test for measuring V617F allele burden.

Systemic Mastocytosis: Molecular Landscape and Implications for Treatment.

Over the past decade, we have witnessed significant advances in the molecular characterization of systemic mastocytosis (SM). This has provided important information for a better understanding of the pathogenesis of the disease but has also practically impacted the way we diagnose and manage it. Advances in molecular testing have run in parallel with advances in therapeutic targeting of constitutive active KIT, the major driver of the disease. Therefore, assessing the molecular landscape in each SM patient is essential for diagnosis, prognosis, treatment, and therapeutic efficacy monitoring. This is facilitated by the routine availability of novel technologies like digital PCR and NGS. This review aims to summarize the pathogenesis of the disease, discuss the value of molecular diagnostic testing and how it should be performed, and provide an overview of present and future therapeutic concepts based on fine molecular characterization of SM patients.

Adverse Prognostic Impact of the KIT D816V Transcriptional Activity in Advanced Systemic Mastocytosis.

In systemic mastocytosis (SM), qualitative and serial quantitative assessment of the KIT D816V mutation is of diagnostic and prognostic relevance. We investigated peripheral blood and bone marrow samples of 161 patients (indolent SM (ISM), n = 40; advanced SM, AdvSM, n = 121) at referral and during follow-up for the KIT D816V variant allele frequency (VAF) at the DNA-level and the KIT D816V expressed allele burden (EAB) at the RNA-level. A round robin test with four participating laboratories revealed an excellent correlation (r > 0.99, R2 > 0.98) between three different DNA-assays. VAF and EAB strongly correlated in ISM (r = 0.91, coefficient of determination, R2 = 0.84) but only to a lesser extent in AdvSM (r = 0.71; R2 = 0.5). However, as compared to an EAB/VAF ratio ≤2 (cohort A, 77/121 patients, 64%) receiver operating characteristic (ROC) analysis identified an EAB/VAF ratio of >2 (cohort B, 44/121 patients, 36%) as predictive for an advanced phenotype and a significantly inferior median survival (3.3 vs. 11.7 years; p = 0.005). In terms of overall survival, Cox-regression analysis was only significant for the EAB/VAF ratio >2 (p = 0.006) but not for VAF or EAB individually. This study demonstrates for the first time that the transcriptional activity of KIT D816V may play an important role in the pathophysiology of SM.

A case of JAK2V617F-positive essential thrombocythemia where allele burden was reduced by a PD-1 inhibitor.

The Janus kinase/signal transducers and activators of transcription signaling pathway induces programmed death ligand-1 (PD-L1) expression. JAK2 mutation at position 617 (JAK2V617) is a frequent driver of myeloproliferative neoplasms (MPN) through PD-L1 expression. Although PD-1 inhibitors should be effective against MPN with JAK2V617F mutation, this has not yet been reported in humans. Thus, we assessed the efficacy of a PD-1 inhibitor in a lung cancer patient with JAK2V617F-positive essential thrombocythemia (ET). A 71-year-old man was diagnosed with ET, and with lung carcinoma 3 years later. After right lobectomy and postoperative chemotherapy, pembrolizumab [a PD-1 inhibitor (200 mg, every 3 weeks)] was initiated for refractory lung carcinoma. Lung cancer progression did not occur for 1.5 years under treatment. Most megakaryocytes were PD-L1-positive, and after pembrolizumab initiation, platelet count remained below 45 × 104/µL without the need for other cytoreductive therapies for ET. The JAK2V617F allele burden gradually decreased from 11.5% at diagnosis to 2.9% after 17 months of pembrolizumab treatment. Other peripheral blood lineages did not decrease, and pembrolizumab treatment was continued without any adverse events. This is the first report demonstrating the effectiveness of pembrolizumab in an MPN patient with JAK2V617F mutation.

Recent Advances in the Molecular Biology of Systemic Mastocytosis: Implications for Diagnosis, Prognosis, and Therapy.

In recent years, molecular characterization and management of patients with systemic mastocytosis (SM) have greatly benefited from the application of advanced technologies. Highly sensitive and accurate assays for KIT D816V mutation detection and quantification have allowed the switch to non-invasive peripheral blood testing for patient screening; allele burden has prognostic implications and may be used to monitor therapeutic efficacy. Progress in genetic profiling of KIT, together with the use of next-generation sequencing panels for the characterization of associated gene mutations, have allowed the stratification of patients into three subgroups differing in terms of pathogenesis and prognosis: i) patients with mast cell-restricted KIT D816V; ii) patients with multilineage KIT D816V-involvement; iii) patients with "multi-mutated disease". Thanks to these findings, new prognostic scoring systems combining clinical and molecular data have been developed. Finally, non-genetic SETD2 histone methyltransferase loss of function has recently been identified in advanced SM. Assessment of SETD2 protein levels and activity might provide prognostic information and has opened new research avenues exploring alternative targeted therapeutic strategies. This review discusses how progress in recent years has rapidly complemented previous knowledge improving the molecular characterization of SM, and how this has the potential to impact on patient diagnosis and management.