Myeloproliferative neoplasms with a low (<5%) CALR mutation allele burden

Abstract Background: CALR mutations were recently identified in a substantial proportion of persons with essential thrombocythemia (ET) and with primary myelofibrosis (PMF) without JAK2V617F. Consequently rapid, sensitive and specific methods to detect and quantify these mutations are needed. Methods: We studied samples from 1088 persons with myeloproliferative neoplasms (MPNs) including 421 JAK2V617F negative subjects with ET, PMF, polycythemia vera (PV), chronic myeloid leukemia (CML) and hyper-eosinophilic syndrome (HES). Detection of CALR exon 9 mutations was done by PCR amplification followed by fragment length analysis and direct sequencing. Dilution assays were used to determine CALR mutant allele burden. Results: We detected CALR mutations in blood and bone marrow samples from 152 subjects with ET and with PMF but not in samples from normal or persons with PV, CML or HES. CALR mutant peaks were distinct from wild-type peaks and dilution experiments indicated a sensitivity level of 0.5-5% for a CALR mutant allele in a wild-type background. Diverse types of mutations were detected including deletions, insertions and complex indels. All mutations were confirmed by direct sequencing. We also used dilution experiments to quantify mutant allele burden. We were able to reproducibly detect mutant allele levels as low 5% (0.5-5%) in a wild-type background. Conclusions: PCR amplification followed by fragment length analysis is a rapid, sensitive and specific method for screening persons with MPNs for CALR mutations, especially those with ET and PV with JAK2V617F and for estimating mutant allele burden. Figure 1. Standard curve for the detection of mutant allele burden. Figure 1. Standard curve for the detection of mutant allele burden. Figure 2. Titration analyses of sensitivity of CALR mutation screening by sequencing and fragment analyses. Figure 2. Titration analyses of sensitivity of CALR mutation screening by sequencing and fragment analyses. Figure 3. Figure 3 Sequencing traces show heterozygous mutation of CALR. Gene scan electropherogram from PCR method and partial sequence of CALR exon 9 from sequencing method (numbering according to GenBank access number: NC_000019.9). A-P: Detected a wild type and 15 CALR mutation types by sequencing and fragment analysis methods. A: wild type, B-I: deletions, H-L: insertions, M-P: complex indels. Figure 3. Figure 3 Sequencing traces show heterozygous mutation of CALR. Gene scan electropherogram from PCR method and partial sequence of CALR exon 9 from sequencing method (numbering according to GenBank access number: NC_000019.9). A-P: Detected a wild type and 15 CALR mutation types by sequencing and fragment analysis methods. A: wild type, B-I: deletions, H-L: insertions, M-P: complex indels. Disclosures No relevant conflicts of interest to declare.

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Pyrosequencing-based quantitative measurement of CALR mutation allele burdens and their clinical implications in patients with myeloproliferative neoplasms

Clinica Chimica Acta ◽

10.1016/j.cca.2018.05.001 ◽

2018 ◽

Vol 483 ◽

pp. 183-191 ◽

Cited By ~ 3

Author(s):

Yejin Oh ◽

Ik-Chan Song ◽

Jimyung Kim ◽

Gye Cheol Kwon ◽

Sun Hoe Koo ◽

...

Keyword(s):

Quantitative Measurement ◽

Myeloproliferative Neoplasms ◽

Clinical Implications ◽

Calr Mutation ◽

Mutation Allele

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Using the Minor Variant Finder software to identify and quantify the allelic burden level of somatic mutations in oncohematologic diseases

Oncohematology ◽

10.17650/1818-8346-2020-15-2-85-91 ◽

2020 ◽

Vol 15 (2) ◽

pp. 85-91

Author(s):

T. N. Subbotina ◽

I. E. Maslyukova ◽

A. A. Faleeva ◽

P. A. Nikolaeva ◽

A. S. Khazieva ◽

...

Keyword(s):

Mutant Allele ◽

Sanger Sequencing ◽

Quantitative Assessment ◽

Somatic Mutations ◽

Myeloproliferative Neoplasms ◽

Mutant Gene ◽

Positive Sample ◽

Allele Burden ◽

Minor Variant ◽

Allelic Burden

Background. There are problems related to both quantitative assessment of an allele burden level of a mutant gene and interpretation of results in DNA samples with the burden level of the mutant allele less than 15–20 %, when using Sanger sequencing for analyzing somatic mutations. Applied Biosystems (USA) has developed new software Minor Variant Finder, which allows determining mutations with the allele burden level from 5 %.The objective: to determine the allele burden level and identification of minor variants of somatic mutations in the ASXL1, JAK2 genes and BCR-ABL oncogene using Minor Variant Finder software in patients with myeloproliferative neoplasms.Materials and methods. The level of mutant allele burden for 15 patients with myeloproliferative neoplasms was determined by the identified mutations using the Minor Variant Finder software, after analysis of point somatic mutations in the ASXL1, JAK2 genes and BCR-ABL oncogene by Sanger sequencing.Results. The allele burden level in all 5 ASXL1-positive samples and BCR-ABL-positive sample was determined as higher than 20 % using the Minor Variant Finder software. The allele burden level in 2 cases was higher than 20 % and in 7 cases lower than 20 %, when we analyzed 9 JAK2-positive samples.Conclusion. Minor Variant Finder software can be used to estimate the allele burden level and to identify minor variants of somatic mutations in the ASXL, JAK2 and BCR-ABL genes.

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Thrombocytapheresis in Patient with Essential Thrombocythemia: A Case Report

Case Reports in Oncology ◽

10.1159/000507651 ◽

2020 ◽

Vol 13 (2) ◽

pp. 675-679

Author(s):

Afra M. Elhassan ◽

Arwa Alsaud ◽

Mohamed A. Yassin ◽

Mahmood Aldapt ◽

Lubna Riaz ◽

...

Keyword(s):

Platelet Count ◽

Case Report ◽

Essential Thrombocythemia ◽

Myeloproliferative Neoplasms ◽

Jak2 V617f ◽

Favorable Outcome ◽

Who Criteria ◽

Icu Admission ◽

Calr Mutation ◽

Clonal Abnormalities

Essential thrombocythemia (ET) is one of the myeloproliferative neoplasms, characterized by persistent thrombocytosis, platelets >450,000/μL, and evident clonal abnormalities like JAK2 V617F, MPL, CALR mutation and not fulfilling WHO criteria for MDS, CML, PV, and IDA. Here we report a 24-year-old female who presented with headache and was found to have thrombocytosis with a platelet count of 2,141 × 103/μL, diagnosed as ET as per WHO criteria 2008; she required ICU admission and thrombocytapheresis with a favorable outcome.

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Influence of JAK2 46/1 haplotype in the natural evolution of JAK2V617F allele burden in patients with myeloproliferative neoplasms

Leukemia Research ◽

10.1016/j.leukres.2011.09.029 ◽

2012 ◽

Vol 36 (3) ◽

pp. 324-326 ◽

Cited By ~ 12

Author(s):

Alberto Alvarez-Larrán ◽

Anna Angona ◽

Luz Martínez-Avilés ◽

Beatriz Bellosillo ◽

Carlos Besses

Keyword(s):

Myeloproliferative Neoplasms ◽

Natural Evolution ◽

Allele Burden ◽

Jak2v617f Allele Burden

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Clinical Implications for Patients with Low JAK2V617F Allele Burden

Blood ◽

10.1182/blood.v120.21.1745.1745 ◽

2012 ◽

Vol 120 (21) ◽

pp. 1745-1745

Author(s):

Marguerite Vignon ◽

Dorota Jeziorowska ◽

Pierre Hirsch ◽

Ollivier Legrand ◽

Nicole Casadevall ◽

...

Keyword(s):

Bone Marrow ◽

Clinical Phenotype ◽

Myeloproliferative Neoplasms ◽

Kinase Domain ◽

Jak2 Mutation ◽

Allele Burden ◽

Vein Thrombosis ◽

Myeloid Neoplasms ◽

Exon 10 ◽

Jak2v617f Allele Burden

Abstract Abstract 1745 In classical Philadelphia-negative myeloproliferative neoplasms (MPN), JAK2V617F is considered as a driver mutation when the threshold of 1% JAK2V617F/JAK2total allele burden is reached. However a lower ratio is sometimes detected with highly sensitive assays. We investigated the clinical significance of such minor clones by describing the characteristics of 27 patients with a low JAK2V617F allele burden ranging from 0.1% to 0.99%. Material and Methods A commercially available quantitative ASO-PCR assay of 0.1% sensitivity (MutaQuant® kit, Ipsogen) was used. Two thousand five hundred consecutive blood samples were sent to our lab for JAK2V617F mutation between 2009 and 2012. Total blood DNA was extracted by an automated standardized procedure (Qiasymphony®, Qiagen). All samples were tested in duplicate. The 27 samples of our cohort were controlled using a second assay of 0.01% sensitivity (Larsen et al, BJH 2007). Thirty samples from healthy donors were also tested. High resolution melting curve (HRM) analysis of JAK2 exon 14 ruled out the possibility of an additional mutation hampering the annealing of a primer. Patients with a known classical MPN clinical phenotype were also tested for JAK2 exons 12–17 (entire pseudo-kinase domain) or for MPL exon 10 depending on the context. Results Laboratory Findings Among the 2500 samples, 735 (29.4%) were positive above 1%, 27 (1.1%) had low JAK2V617F allele burden ranging from 0.12 to 0.99%. The patient with the lowest ratio (0.12%) was not confirmed by the second assay and therefore was excluded from the study. This allowed the median to settle at 0.40%. No associated mutations were found in the JAK2 pseudo-kinase domain in patients with polycythemia vera (PV) and in MPL exon 10 in patients with essential thrombocytosis (ET) and primary myelofibrosis (PMF). Healthy patients were all tested JAK2V617F negative. Clinical Aspects The cohort included 19 men and 7 women ranging from 28 to 95 years of age (median 63 years old). Two patients had secondary acute myeloid leukaemia following JAK2V617F positive MPN indicating the presence of residual JAK2V617F cells and the negativity of the myeloblastic population. Thirteen patients (50%) had a classical MPN with a median ratio of 0.36%: 7 ET, 5 PV and 1 PMF according to WHO 2008 criteria. However a bone marrow biopsy was available for only two patients (1 ET, 1 PMF). None of them had received pegylated interferon alpha-2a. Four patients had a prior history of thrombosis: two strokes, one pulmonary embolism, two portal vein thrombosis (PVT). For one PV patient, a 6 months follow-up blood and bone marrow sample confirmed a low allele burden in the same range (0.4%) and in vitro Epo-independant erythroid colonies were observed. Five patients had other chronic myeloid neoplasms (two myelodysplastic/myeloproliferative neoplasms, one chronic eosinophilic leukaemia, one chronic myeloid leukaemia, one refractory anaemia with ring sideroblasts). Among these five, four had an abnormal karyotype. We did not observe any thrombotic event in these patients. We cannot conclude on hematological diagnosis for the last six patients: four patients were screened for JAK2 mutation because of PVT. One patient had chronic polycythemia in a context of alcohol and tobacco abuse. One patient had homozygous hemochromatosis with a normal haemoglobin level in spite of repeated phlebotomies. Discussion In this single centre study low JAK2V617F allele burden represented 1% of all samples sent for JAK2V617F study and 3.5% of JAK2V617F positive patients. Seventeen patients (65%) had classical MPN or splanchnic vein thrombosis. To our knowledge PV patients with such low JAK2V617F allele burden have not been reported in the absence of associated JAK2 pseudo-kinase domain mutation. A larger screen for cooperating mutations responsible for the PV phenotype is under process. In the context of other chronic myeloid neoplasms, the JAK2V617F mutation is thought to belong to a more complex clonal architecture mostly implicating chromatin remodeling genes. Here, the presence of a JAK2 mutation could argue in favour of clonal haematopoiesis. In conclusion the clinical phenotype of low JAK2V617F patients overlaps with classical JAK2V617F MPN. The technical implications might be challenging for molecular diagnostic platforms. More data are needed to further characterize these patients. Disclosures: No relevant conflicts of interest to declare.

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Correlations Between TET2 Mutation and Clinicohematologic Parameters in Myeloproliferative Neoplasms

Blood ◽

10.1182/blood.v120.21.1455.1455 ◽

2012 ◽

Vol 120 (21) ◽

pp. 1455-1455

Author(s):

Jung Sook Ha ◽

Jae Hee Lee ◽

Sung Gyun Park ◽

Nam Hee Ryoo ◽

Dong Suk Jeon ◽

...

Keyword(s):

Conflicts Of Interest ◽

Myeloproliferative Neoplasms ◽

Direct Sequencing ◽

Jak2 V617f ◽

Putative Tumor Suppressor Gene ◽

Allele Burden ◽

Frame Shift ◽

Mutational Status ◽

Tet2 Mutation ◽

Disease Specific

Abstract Abstract 1455 Background: Since the acquired somatic mutation, JAK2 V617F, was discovered as a first molecular marker of myeloproliferative neoplasms (MPN), and it has been detected variably in each MPN subtypes. However, JAK2 V617F does not found in all of MPN cases and not necessarily specific to a particular clinicpathologic entity. Recently, mutation of the putative tumor suppressor gene, Ten-Eleven-Translocation-2(TET2), has been identified in MPN patients. However, the frequency of TET2 mutation or its relationship with JAK2 V617F mutation or pathologic function in MPN has not been concluded, yet. The aim of our study was to evaluate the frequency of TET2 in MPN patients, and whether there is any correlation of TET2 mutation with JAK2V617F mutation or the clinicohematologic parameters. Materials and Methods: Total 99 adult MPN patients (18 PV, 62 ET, 11 PMF and 8 MPN unclassified) whose bone marrow cells had been stored from 2007 to 2010 at point of first diagnosis were included in this study. Hematological diagnoses and subtyping were reconfirmed according to the 2008 WHO classification and clinicohematologic datas were collected from patient records. Direct sequencing for TET2(exon3–11) and JAK2 (exons 12 and 14) were performed using an ABI 3730XL DNA analyzer. The JAK2V617F allele burdens were determined by pyrosequencing for samples available and MPL was analyzed by allele-specific PCR. Results: The overall TET2 mutational frequency was 12.1%, and disease-specific mutational frequencies were 22.2% in PV, 9.7% in ET and 18.2% in PMF. The found mutations included 11 mutations, 7 frame-shift (p.Lys95AsnfsX18, p.Gln967AsnfsX40, p.Lys1022GlufsX4, p.Asp1314MetfsX49, p.Gln1534AlafsX43, p.Tyr1618LeufsX4, p.Leu1609GlufsX45), 1 nonsense (p.Gly1735X), 1 missense (Q599R) and 2 splicing mutations (c.3409+1G>T, c.4044+2insT). Those mutations most frequently involved exon 3(four mutations) and exon 11(four mutaions), and rarely intron 3, intron 8 and exon 7. None of the mutations were associated with a karyotypically apparent 4q24 rearrangement. All patients were also screened for JAK2 V617F, and the overall JAK2 V617F positive rate was 68%(94.4% in PV, 69.4% in ET, 45.5% in PMF and 37.5% in MPN, unclassified). All TET2 mutations occurred in JAK2 V617F positive cases. JAK2 exon12 mutation was not found in all patients. MPL W515L was found in one ET patient who also carried JAK2V617F, but not TET2 mutation. Information on JAK2 V617F allele burden was available in 78 patients. Considering all 99 patients, the patient age, hematologic indexes (leukocyte count, neutrophil fraction, lymphocyte fraction, monocyte fraction, Hb, Hct and platelet count), the frequency of organomegaly, marrow fibrosis or thrombotic/hemorrhagic complications were not different according to carrying TET2 mutation. However, TET2 mutation was more frequently found in JAK2 V617F carriers than non-carriers (P=0.008), but JAK2 V617F allele burden did not correlated with the presence of mutant TET2. When analysis was performed for each PV, ET, and PMF (no TET2 mutation in MPN-unclassifiable patients), correlation between TET2 and JAK2 V617F mutational status was not found in each subtypes (P=0.078 in PV, P=0.099 in ET and P=0.182 in PMF). However, the JAK2 V617F allele burden was significantly higher in PMF harboring TET2 mutation than PMF patients did not (88.0 ± 4.3% vs 19.1 ± 28.7%, P=0.034). In statistical analysis for the correlations of clinicohematologic parameters with TET2 mutation in each PV, ET and PMF patients, only a few statistically significant results were identified. The presence of TET2 mutation was correlated with high Hct in PMF (47.4 ± 5.4 vs 25.5 ± 6.2, P=0.037), and TET2 positive ET patients showed relatively higher frequency of organomegaly compared to ET patients without TET2 mutation (50% vs 19.6%, P=0.018). Conclusions: The overall and disease-specific frequencies of TET2 mutation in our study are similar with previous studies, and frame-shift mutation is the most frequent mutation type. There is no specific relationship between JAK2 V617F and TET2 mutation occurrence, but TET2 mutant PMF has higher JAK2 V617F allele burden than non-mutant. TET2 mutation is also associated with a higher Hct in PMF and higher frequency of organomegaly in ET. Larger scale studies involving more MPN patients are needed. Disclosures: No relevant conflicts of interest to declare.

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The Role Of Deregulated HMGA2 Expression With Promoter Methylation Of p16 In Myeloproliferative Neoplasms

Blood ◽

10.1182/blood.v122.21.1606.1606 ◽

2013 ◽

Vol 122 (21) ◽

pp. 1606-1606

Author(s):

Kayo Shirado Harada ◽

Kazuhiko Ikeda ◽

Kazuei Ogawa ◽

Hideyoshi Noji ◽

Hideo Kimura ◽

...

Keyword(s):

Promoter Methylation ◽

Blood Cells ◽

Conflicts Of Interest ◽

Myeloproliferative Neoplasms ◽

Expression Levels ◽

Allele Burden ◽

Polycomb Group Genes ◽

Micro Rnas ◽

Jak2v617f Allele Burden

Abstract Myeloproliferative Neoplasms (MPNs) are characterized by clonal proliferative hematopoiesis with increased mature blood cells. The signal-activating mutations such as JAK2V617F increase blood cells, but it remains uncertain how an abnormal hematopoietic cell clone expands in MPNs. We have recently showed that overexpression of the high mobility group AT-hook 2 (HMGA2) causes proliferative hematopoiesis with providing a clonal growth advantage to hematopoietic cells in mice (Ikeda et al, Blood, 2011), suggesting the possibility that HMGA2 contributes to the pathogenesis of MPNs. However, since only a few studies have evaluated expression of HMGA2 mRNA in patients with MPNs, the role of HMGA2 in the pathogenesis of MPNs is yet unclear. MPNs also show mutations in epigenetic modifiers involving DNA methylation such as polycomb group genes (PcG) and aberrant expressions of micro RNAs (miRNA) that negatively regulate expressions of targeted genes. Interestingly, deficiency in either PcG-related BMI1 (Oguro et al, J Exp Med, 2012) or let-7-family miRNA (Mayr et al, Science, 2007) causes deregulation of HMGA2 expression, leading to its oncogenic activity in part by negatively regulating tumor suppressor p16. Thus, in this study, to clarify the role of HMGA2 in MPNs, we investigated expression of HMGA2 mRNA in peripheral granulocytes of 56 patients with MPNs including 23 polycythemia vera (PV), 26 essential thrombocythemia (ET) and 7 primary myelofibrosis (PMF) along with clinical findings, JAK2V617F allele burden, expressions of BMI1 mRNA and let-7-family miRNAs, and promoter methylation of p16. Quantitative RT-PCR (qPCR) showed significantly higher expression of HMGA2 mRNA relative to internal control HPRT1 mRNA in PMF (mean ± SD; 31.7 ± 42.8, p<0.01), but not PV (15.7 ± 53.2) or ET (2.14 ± 7.70), compared with 12 healthy volunteers (HV; 0.431 ± 0.366). In addition, deregulated HMGA2 expression (>1.2), which was determined as relative expression level above mean + 2SD of HMGA2 mRNA in 12 HV, was most frequently detected in patients with PMF [7/7 (100%)] (p<0.01), compared with PV [5/23 (21.7%)] and ET [6/26 (23.1%)]. We also found a significant positive correlation in expression levels of HMGA2 mRNA with serum LDH values (r=0.531, p<0.01) rather than JAK2V617F allele burden (r=0.25, p=0.08). These data suggested that expression of HMGA2 mRNA independently correlated with disease phenotype and status in MPNs. We next explored the cause of deregulated expression of HMGA2 mRNA and found lower expression of let-7a (0.19 ± 0.13 vs. 0.42 ± 0.39, p=0.04) and -7c (0.57 ± 0.60 vs. 1.14 ± 0.94, p=0.06) rather than -7b (p=0.2) by qPCR, in patients with deregulated expression of HMGA2 mRNA compared with other patients. However, HMGA2-involved chromosomal abnormality in 12q13-15 was not detected in any patient, and there was no difference in expression of BMI1 mRNA between patients with deregulated expression of HMGA2 mRNA and other patients. Thus, decreased expression of let-7 miRNAs might contribute to deregulated expression of HMGA2 mRNA in MPNs. Finally, we investigated correlation of deregulated expression of HMGA2 mRNA with promoter methylation of p16. Methylation-specific PCR assay detected promoter methylation of p16 in 17/56 (30.4%) patients with MPNs. Strikingly, patients with deregulated expression of HMGA2 mRNA significantly more often showed promoter methylation of p16 compared with other patients [10/18 (55.6%) vs. 7/38 (18.4%), p<0.01]. Furthermore, patients with promoter methylation of p16 showed higher expression levels of HMGA2 mRNA than patients without the methylation, especially in patients with PMF (2.33 ± 0.90 vs. 70.9 ± 38.3, p=0.01). In conclusion, deregulated expression of HMGA2 in association with decreased expression of let-7 miRNAs may play a crucial role in the pathogenesis of MPNs possibly through p16. Disclosures: No relevant conflicts of interest to declare.

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Latent Myeloproliferative Neoplasm May Underlie Retinal Vein Occlusion In Older Patients

Blood ◽

10.1182/blood.v122.21.5242.5242 ◽

2013 ◽

Vol 122 (21) ◽

pp. 5242-5242

Author(s):

Katerina Zoi ◽

Christine Zoi ◽

Andreas Giannopoulos ◽

Argyri Gialeraki ◽

Kassiani Giannaki ◽

...

Keyword(s):

Retinal Vein Occlusion ◽

Older Patients ◽

Conflicts Of Interest ◽

Myeloproliferative Neoplasms ◽

Significant Risk ◽

Jak2v617f Mutation ◽

Thrombotic Complication ◽

Allele Burden ◽

Vein Occlusion ◽

Increased Risk

Abstract Background Myeloproliferative neoplasms (MPNs) have been associated with a high incidence of thrombosis and bleeding episodes, which significantly contribute to disease-related morbidity and mortality. Clinical data indicate an association of the JAK2V617F mutation, seen in nearly all polycythemia vera (PV) cases and almost 60% of those with essential thrombocythemia (ET) and myelofibrosis (MF). The mutation is also seen in 37% of patients with in splachnic vein thrombosis (SVT) and its presence was associated with an increased risk for SVT. However, the prevalence of JAK2V617 seems to be low in patients with other thromboembolic events in unusual sites such as cerebral sinus, upper limb deep venous thrombosis (DVT). Additionally, activating mutations of MPL gene, seen in 3% of ET and 5% of MF patients, are considered as a significant risk factor for microvessel disturbances and have been associated with an increased risk of arterial thrombosis. Retinal vein occlusion (RVO) is a thrombotic complication in an uncommon site that may result in sight threatening disease. In this study we investigated the prevalence of JAK2V617F and MPLW515L/K mutations in a prospectively assembled cohort of patients with RVO, hypothesizing that some cases may be associated with an underlying undiagnosed MPN. Patients and Methods We studied 52 (23 males and 29 females) consecutive patients with no evidence of an underlying MPN who had been diagnosed with RVO confirmed with fluorangiography from January 2007 to September 2011. The mean age was 70 years (range: 49-85) Twenty eight patients (53.8%) presented with central RVO and 24 patients with branched RVO (46.5%). DNA was extracted from peripheral blood samples by standard procedures. The JAK2V617F mutation was detected using a tetra-primer amplification refractory mutation system (ARMS) polymerase chain reaction (PCR) assay with a sensitivity of 1% and the allele burden was estimated with a semi-quantitative method. MPLW515L/K were detected using allele-specific PCR (AS-PCR) assays with a sensitivity of 1%. Results Overall, MPN associated mutations were detected in 5/52 cases. JAK2V617F was detected in 2/52 cases (3.8%; 95%CI-1.4%-9%), while MPL exon 10 mutations were detected in 3/52 (5.7%; 95%CI-0.6%-12%). The JAK2V617F allele burden in the two positive patients was 45% and 52% respectively. Both patients who carried the JAK2V617F mutation were female. The first patient had been already diagnosed with ET according to the WHO criteria at the time of RVO screening. She was receiving hydroxyurea and aspirin and her platelet count was normal. The second patient who also carried the JAK2V617F mutation had a PLT count of 850.000/μl at the time of screening and was diagnosed with ET within the 3 following months. The patients with MPL mutations presented with normal blood counts. Conclusions Our findings indicate that a latent MPN could underlie RVO even in the absence of conventional diagnostic criteria. Our results represent the first report that MPL mutations could underlie RVO cases and suggest that routine screening of RVO cases for MPN mutations may be useful, especially in older patients. Disclosures: No relevant conflicts of interest to declare.

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