JAK-2 V617F Mutation in Endothelial Cells of Patients with Atherosclerotic Carotid Disease.

Objective: It has been shown that clonal mutations occur in hematopoietic stem cells with advancing age and increase the risk of death due to atherosclerotic vascular diseases, just like in myeloproliferative neoplasms. It is known that endothelial cells (EC) and hematopoietic stem cells develop from a common stem cell called hemangioblast in early embryonic period. However, the presence of hemangioblast in the postnatal period is controversial. In this study, JAK2 gene variants was examined in patients with atherosclerotic carotid disease and without any hematological malignancy. Materials and Methods: Ten consecutive patients (8 men and 2 women) with symptomatic atherosclerotic carotid stenosis were included in this study. EC (CD31+CD45-) were separated from tissue samples taken by carotid endarterectomy. JAK2 variants was examined in EC, peripheral blood mononuclear cells and oral epithelial cells of the patients with next generation sequencing. Results: The median age of the patients was 74 (58-80) and the median BMI was 24,44 (18,42-30,85) kg/m2. Smoking history was present in 50%, hypertension in 80%, diabetes in 70%, and ischemic heart disease in 70% of the patients. JAK2V617F mutation was detected in peripheral blood mononuclear cells in three out of 10 patients, two of them also had JAK2V617F mutation in their EC. JAK2V617F mutation was not found in oral epithelial cells in any of the patients. Conclusion: In this study, for the first time in the literature, we showed that JAK2V617F mutation was found somatically in both peripheral blood cells and EC in patients with atherosclerosis. This finding may support that EC and hematopoietic cells originate from a common clone or that the somatic mutation can be transmitted to EC by other mechanisms. Examining the molecular and functional changes caused by JAK2V617F mutation in EC may help open a new avenue for treating atherosclerosis.

Impact of CALR and JAK2V617F Mutations on Clinical Course and Disease Outcomes in Essential Thrombocythemia: A Multicenter Retrospective Study in Turkish Patients.

Objective: In this study, we investigated the effects of calreticulin (CALR) and JAK2V617F mutational status on clinical course and disease outcomes in Turkish patients with essential thrombocythemia (ET). Materials and Methods: Seventeen centers from Türkiye participated in the study and CALR- and JAK2V617F-mutated ET patients were evaluated retrospectively. Results: A total of 302 patients were included, of whom 203 (67.2%) and 99 (32.8%) were JAK2V617F- and CALR-positive, respectively. CALR-mutated patients were significantly younger (51 years vs. 57.5 years, p=0.03), with higher median platelet counts (987x109/L vs. 709x109/L, p<0.001) and lower median hemoglobin levels (13.1 g/dL vs. 14.1 g/dL, p<0.001) compared to JAK2V617F-mutated patients. Thromboembolic events (TEEs) occurred in 54 patients (17.9%), 77.8% of which were arterial. Compared to CALR mutation, JAK2V617F was associated with a higher risk of thrombosis (8.1% vs. 22.7%, p=0.002). Rates of transformation to myelofibrosis (MF) and leukemia were 4% and 0.7%, respectively, and these rates were comparable between JAK2V617F- and CALR-mutated cases. The estimated overall survival (OS) and MF-free survival of the entire cohort were 265.1 months and 235.7 months, respectively. OS and MF-free survival durations were similar between JAK2V617F- and CALR-mutated patients. Thrombosis-free survival (TFS) was superior in CALR-mutated patients compared to JAK2V617F-positive patients (5-year TFS: 90% vs. 71%, respectively; p=0.001). Age at diagnosis was an independent factor affecting the incidence of TEEs. Conclusion: In our ET cohort, CALR mutations resulted in higher platelet counts and lower hemoglobin levels than JAK2V617F and were associated with younger age at diagnosis. JAK2V617F was strongly associated with thrombosis and worse TFS. Hydroxyurea was the most preferred cytoreductive agent for patients with high thrombosis risk.

Immune Thrombotic Thrombocytopenic Purpura in Elderly Patients: The Roles of PLASMIC and French Scores

Objective: In recent years, new developments have been incorporated into daily practice in the management of immune thrombotic thrombocytopenic purpura (iTTP). In particular, clinical scoring systems could help clinicians with clinical decision-making and early recognition. However, older patients frequently present with more organ involvement and in unusual ways. The ways in which age could affect these clinical prediction scoring systems remain unclear. We evaluated the use of PLASMIC and French scores in patients over 60 years of age. Materials and Methods: We performed a retrospective cross-sectional analysis of patients over 60 years of age with a presumptive diagnosis of iTTP between 2014 and 2022 at 10 centers. We calculated PLASMIC and French scores and compared our data with a single-center analysis of younger patients presenting with thrombotic microangiopathy. Results: Our study included 30 patients over 60 years of age and a control group of 28 patients younger than 60 years. The diagnostic sensitivity and specificity of a French score of ≥1 were lower in older patients compared to the control group (78.9% vs. 100% and 18.2% vs. 57.1%, respectively). The diagnostic sensitivity and specificity of a PLASMIC score of ≥5 were 100% vs. 95% and 27.3% vs. 100% for the study group and control group, respectively. Our study showed a higher mortality rate in older patients compared to the control group (30% vs. 7.1%, p=0.043). Conclusion: For a limited number of patients (n=6), our results showed that rituximab can reduce mortality. Given that the reliability of clinical prediction scores for iTTP in older patients may be lower, more caution must be undertaken in interpreting their results.

The 2023 ACR/EULAR Antiphospholipid Syndrome Classification Criteria.

OBJECTIVE: To develop new antiphospholipid syndrome (APS) classification criteria with high specificity for use in observational studies and trials, jointly supported by the American College of Rheumatology (ACR) and EULAR. METHODS: This international multidisciplinary initiative included 4 phases: 1) Phase I, criteria generation by surveys and literature review; 2) Phase II, criteria reduction by modified Delphi and nominal group technique exercises; 3) Phase III, criteria definition, further reduction with the guidance of real-world patient scenarios, and weighting via consensus-based multicriteria decision analysis, and threshold identification; and 4) Phase IV, validation using independent adjudicators' consensus as the gold standard. RESULTS: The 2023 ACR/EULAR APS classification criteria include an entry criterion of at least one positive antiphospholipid antibody (aPL) test within 3 years of identification of an aPL-associated clinical criterion, followed by additive weighted criteria (score range 1-7 points each) clustered into 6 clinical domains (macrovascular venous thromboembolism, macrovascular arterial thrombosis, microvascular, obstetric, cardiac valve, and hematologic) and 2 laboratory domains (lupus anticoagulant functional coagulation assays, and solid-phase enzyme-linked immunosorbent assays for IgG/IgM anticardiolipin and/or IgG/IgM anti-ß2 -glycoprotein I antibodies). Patients accumulating at least 3 points each from the clinical and laboratory domains are classified as having APS. In the validation cohort, the new APS criteria versus the 2006 revised Sapporo classification criteria had a specificity of 99% versus 86%, and a sensitivity of 84% versus 99%. CONCLUSION: These new ACR/EULAR APS classification criteria were developed using rigorous methodology with multidisciplinary international input. Hierarchically clustered, weighted, and risk-stratified criteria reflect the current thinking about APS, providing high specificity and a strong foundation for future APS research.

2023 ACR/EULAR antiphospholipid syndrome classification criteria.

OBJECTIVE: To develop new antiphospholipid syndrome (APS) classification criteria with high specificity for use in observational studies and trials, jointly supported by the American College of Rheumatology (ACR) and EULAR. METHODS: This international multidisciplinary initiative included four phases: (1) Phase I, criteria generation by surveys and literature review; (2) Phase II, criteria reduction by modified Delphi and nominal group technique exercises; (3) Phase III, criteria definition, further reduction with the guidance of real-world patient scenarios, and weighting via consensus-based multicriteria decision analysis, and threshold identification; and (4) Phase IV, validation using independent adjudicators' consensus as the gold standard. RESULTS: The 2023 ACR/EULAR APS classification criteria include an entry criterion of at least one positive antiphospholipid antibody (aPL) test within 3 years of identification of an aPL-associated clinical criterion, followed by additive weighted criteria (score range 1-7 points each) clustered into six clinical domains (macrovascular venous thromboembolism, macrovascular arterial thrombosis, microvascular, obstetric, cardiac valve, and hematologic) and two laboratory domains (lupus anticoagulant functional coagulation assays, and solid-phase enzyme-linked immunosorbent assays for IgG/IgM anticardiolipin and/or IgG/IgM anti-ß2-glycoprotein I antibodies). Patients accumulating at least three points each from the clinical and laboratory domains are classified as having APS. In the validation cohort, the new APS criteria vs the 2006 revised Sapporo classification criteria had a specificity of 99% vs 86%, and a sensitivity of 84% vs 99%. CONCLUSION: These new ACR/EULAR APS classification criteria were developed using rigorous methodology with multidisciplinary international input. Hierarchically clustered, weighted, and risk-stratified criteria reflect the current thinking about APS, providing high specificity and a strong foundation for future APS research.

Different perspectives on translational genomics in personalized medicine

Personalized medicine is a relatively new and interesting concept in the medical and healthcare industries. New approaches in current research have supported the search for biomarkers, based on the genomic, epigenomic and proteomic profile of individuals, using new technological tools. This perspective involves the potential to determine optimal medical interventions and provide the optimal benefit-risk balance for treatment, whilst it also takes a patient's personal situation into consideration. Translational genomics, a subfield of personalized medicine, is changing medical practice, by facilitating clinical or non-clinical screening tests, informing diagnoses and therapeutics, and routinely offering personalized health-risk assessments and personalized treatments. Further research into translational genomics will play a critical role in creating a new approach to cancer, pharmacogenomics, and women's health. Our current knowledge may be used to develop new solutions that can be used to minimize, improve, manage, and delay the symptoms of diseases in real-time and maintain a healthy lifestyle. In this review, we define and discuss the current status of translational genomics in some special areas including integration into research and health care.

Comparison of Response to Rituximab Therapy in Adults with Refractory Symptomatic Immune Thrombocytopenia According to the Presence of Accessory Spleen.

No data exist for the association between the presence of accessory spleen after splenectomy and response to rituximab in immune thrombocytopenia (ITP). We investigated the relationship between accessory spleen presence and rituximab response in splenectomized ITP patients. Fifteen chronic refractory ITP patients were included. Four weekly doses of rituximab 375 mg/m2 were administered. All patients had undergone splenectomy before rituximab administration. Accessory spleen was detected in 5 of 15 patients (33.3%). Median age at diagnosis was significantly higher in patients with accessory spleen than those without accessory spleen (40 (range 25-68 years) and 26 (range 7-40 years), respectively; p = 0.049). There was a trend for older age at time of rituximab initiation in patients with accessory spleen compared to the other group (median 51 (range 43-75 years) and 42.5 (range 30-60 years), respectively; p = 0.066). Median follow-up duration was 96 months (range 40-98). We demonstrated a significant correlation between accessory spleen presence and older age. Accessory spleen presence correlated with higher platelet and WBC counts. We showed good inverse correlation between presence of accessory spleen and time to early response (ER) to rituximab while the rate of early response (ER), late response (LR), sustained response (SR) and overall response (OR) did not differ with respect to the presence of acessory spleen.

Description of damage in different clusters of patients with antiphospholipid syndrome.

OBJECTIVE: To identify the different clinical phenotypes of antiphospholipid syndrome (APS) by using cluster analysis and describe cumulative damage of disease clusters. METHODS: This retrospective study includes patients with APS (±systemic lupus erythematosus (SLE)). Two-step cluster analysis was applied by considering clinical data. Damage was calculated for all patients by applying damage index for APS (DIAPS). RESULTS: A total of 237 patients (198 females; median age of 43 years; median follow-up of 9.5 years) were classified into four clusters. Cluster 1 (n = 74) consisted of older patients with arterial-predominant thrombosis, livedo reticularis, and increased cardiovascular risk; cluster 2 (n = 70) of SLE+APS patients with thrombocytopenia and heart valve disease; cluster 3 (n = 59) of patients with venous-predominant thrombosis, less extra-criteria manifestations; and cluster 4 (n = 34) of patients with only pregnancy morbidity with lower frequency of extra-criteria features and cardiovascular risk. Patients with SLE+APS (n = 123) had the highest mean DIAPS. Regarding clusters, 1 and 2 had high cumulative damage. While cumulative survival rates of clusters did not differ, cluster 2 and 3 had lower survival rates at further years. There was no correlation between DIAPS and mortality. CONCLUSION: SLE+APS patients with extra-criteria manifestations and older APS patients with arterial thrombosis and increased cardiovascular risk have higher cumulative damage. Effective treatment of SLE disease activity and control of cardiovascular risk may help to reduce cumulative damage in these patients.

A Novel ATP6V0A2 Mutation Causing Recessive Cutis Laxa with Unusual Manifestations of Bleeding Diathesis and Defective Wound Healing

Objective: Autosomal recessive cutis laxa type IIA (ARCL2A) is a rare congenital disorder characterized by loose and elastic skin, growth and developmental delay, and skeletal anomalies. It is caused by biallelic mutations in ATP6V0A2. Those mutations lead to increased pH in secretory vesicles and thereby to impaired glycosyltransferase activity and organelle trafficking. We aimed to identify the genetic and molecular cause of the unexpected hematological findings in a Turkish family. Materials and Methods: We performed clinical, genetic, and histological analyses of a consanguineous family afflicted with wrinkled and loose skin, microcephaly, intellectual disability, cleft lip and palate, downslanting palpebral fissures, ectopia lentis, bleeding diathesis, and defective wound healing. Results: Linkage analysis using SNP genotype data yielded a maximal multipoint logarithm of odds score of 2.59 at 12q24.21-24.32. Exome sequence analysis for the proband led to the identification of novel homozygous frameshift c.2085_2088del (p.(Ser695Argfs*12)) in ATP6V0A2, within the linked region, in the two affected siblings. Conclusion: Our patients do not have gross structural brain defects besides microcephaly, strabismus, myopia, and growth or developmental delay. Large platelets were observed in the patients and unusual electron-dense intracytoplasmic inclusions in fibroblasts and epidermal basal cells were observed in both affected and unaffected family members. The patients do not have any genetic defect in the VWF gene but von Willebrand factor activity to antigen ratios were low. Clinical findings of bleeding diathesis and defective wound healing have not been reported in ARCL2A and hence our findings expand the phenotypic spectrum of the disease.

The association between P selectin glycoprotein ligand 1 gene variable number of tandem repeats polymorphism and risk of thrombosis in Behçet's disease.

OBJECTIVES: Behçet's disease (BD) has been recognized as an unclassified type of vasculitis with an accompanying tendency to thrombosis. No disease-specific pathology has been demonstrated so far to explain the prothrombotic state, and this predisposition is considered to be associated with endothelial activation/dysfunction. P-selectin glycoprotein ligand-1 (PSGL-1) variable number of tandem repeat (VNTR) polymorphism has an impact on the protein length, and heterozygosity affect of the PSGL-1 to P-selectin interaction, which has been found to be associated with an increased risk of thrombosis in patients with antiphospholipid syndrome. We aimed to analyze the association of PSGL-1 gene polymorphism, in a group of BD patients with and without thrombosis. METHODS: The study group consisted of 136 BD patients (112 male, 24 female) with thrombosis, 120 BD patients without thrombosis (54 male, 66 female) during at least 5 years disease course, and 190 healthy controls (103 male, 87 female) All patients fulfilled the International Study Group criteria for classification of BD. Genotyping for the PSGL-1 gene exon 2 VNTR polymorphism was carried out with the amplification of genomic DNA and running of the polymerase chain reaction product on agarose gel electrophoresis. RESULTS: The frequency of heterozygous genotypes (AB+AC+BC) was greater in BD patients with thrombosis compared to BD patients without thrombosis (33.1% vs. 20.8%, P = 0.028, odds ratio = 1.85). However, the increased frequency of heterozygous genotypes in BD patients with thrombosis did not reach a statistically significant level compared to healthy controls (33.1% vs. 32.6%). CONCLUSIONS: PSGL-1 VNTR polymorphism may have limited contribution to the thrombotic tendency in patients with BD.

Rituximab Therapy in Adults with Refractory Symptomatic Immune Thrombocytopenia: Long-Term Follow-Up of 15 Cases.

OBJECTIVE: This paper prospectively evaluates the long-term follow-up [mean ± standard deviation (SD) duration: 89.7±19.4 months] data of 15 patients (13 females and 2 males) with refractory symptomatic immune thrombocytopenia (ITP) treated with rituximab. MATERIALS AND METHODS: Rituximab was administered at 375 mg/m2 weekly for a total of 4 doses. Complete response (CR) was defined as a platelet count of ≥100,000/mm3 and partial response (PR) as a platelet count of ≥30,000/mm3 but less than 100,000/mm3. Early response (ER) and late response (LR) were defined as response within 42 days and after 42 days of initiation of rituximab therapy, respectively. Sustained response (SR) was defined as response lasting for at least 6 months. RESULTS: Mean age (±SD) at the start of rituximab was 46.6±11.3 years. Mean platelet count (±SD) prior to rituximab treatment was 17,400±8878/mm3. The mean time (±SD) between rituximab therapy and response to rituximab in early responders and late responders was 1.8±1.3 weeks and 10±2.8 weeks, respectively. Mean durations (±SD) of ER and LR were 51±47.2 months and 6±4.2 months, respectively. Seven of the 15 patients (46.7%) showed an initial response to rituximab (5 ER and 2 LR). The rate of SR over 6 months was 26.7% (4/15). Among the responders to rituximab, 3 (3/7, 42.9%) maintained their response 1 year after rituximab treatment and 2 (2/7, 28.6%) had ongoing response 5 years after initiation of rituximab. Two of the 7 patients (28.6%) still maintained their response 98 months after initiation of rituximab. All 5 initial responders with subsequent relapse achieved response from subsequent treatment modalities (3 CR, 2 PR). CONCLUSION: Our data confirm, over a long period of observation, that rituximab is safe and effective in the management of patients with chronic refractory primary ITP.

Lupus anticoagulant, thrombosis, and death.

In this issue of Blood, Gebhart et al report a prospective observational cohort study evaluating 151 patients with persistently positive lupus anticoagulant (LA) for a median period of 8.2 years. They observed increased mortality in LA-positive patients, mainly due to new thrombotic events.

Selective Spleen Scintigraphy in the Evaluation of Accessory Spleen/Splenosis in Splenectomized/Nonsplenectomized Patients and the Contribution of SPECT Imaging.

OBJECTIVE: We aimed to evaluate the results of selective spleen scintigraphy (SSS) and contribution of SPECT imaging to planar imaging in splenectomized and nonsplenectomized patients. METHODS: We retrospectively examined 112 SSSs of 96 patients. The patients were divided into two groups as splenectomized group (SP) and non-splenectomized group (NSP). The findings were evaluated by comparing the results of surgery,computerized tomography (CT), ultrasonography (USG) and magnetic resonance imaging (MRI). In addition, whether or not differences existed between the results of SPECT and planar imaging was determined. RESULTS: Of 66 scintigraphies performed in the NSP group, 3 (5%) had positive, 3 (5%) had suspicious and 60 (90%) had negative results. In the NSP group, 28 patients underwent surgery and 12 accessory spleens were removed. Only 3 of these tissues were detected by scintigraphy. Of 46 patients in the SP group, 26 (57%) had positive findings whereas 20 (43%) had negative scintigraphies. Twelve accessory spleens/splenosis were removed surgically in 10 patients with a positive SSS in the SP group. There were no false positive results in both groups of patients who underwent surgery. There was no significant difference between the results of SSS, USG and CT. Of 39 patients to whom SPECT were performed, 10 had positive results both with planar and SPECT imaging. On the other hand, 26 patients, 3 of whom had suspected findings in SPECT images, demonstrated negative results when evaluated with both imaging methods. Remaining 3 were considered suspicious by only SPECT images for the hilar area. CONCLUSION: SSS has high specificity in the detection of accessory spleens/splenosis. The sensitivity of SSS is low in the NSP group,but higher in the SP group. There is no contribution of SPECT imaging to planar imaging.

The significance and management of thrombocytopenia in antiphospholipid syndrome.

The association between antiphospholipid antibodies (aPL) and clinical problems goes beyond what is stated in the antiphospholipid syndrome (APS) classification criteria, namely thrombosis and pregnancy morbidity, and thrombocytopenia is the most common non-criteria hematologic manifestation of aPL with a frequency ranging from 20 to 50 %. Thrombocytopenia is rarely severe, and hemorrhage is far less common than thrombosis. However, when anticoagulation is considered, it may constitute a clinical problem with increased bleeding risk. Furthermore, thrombocytopenia represents a risk factor for thrombosis in aPL-positive patients. Therefore, it is important to understand the pathogenesis and the clinical associations of thrombocytopenia to build the right medical approach in aPL-positive patients. In this paper, we review the literature on aPL/APS-associated thrombocytopenia and briefly discuss the other conditions that can result in thrombocytopenia as they have commonalities with APS and their recognition is important to establish the most appropriate treatment strategy.

The Relationship between P-Selectin Polymorphisms and Thrombosis in Antiphospholipid Syndrome: A Pilot Case-Control Study.

OBJECTIVE: The selectins are cell adhesion molecules that mediate the interactions among leukocytes, activated platelets, and endothelial cells. We aimed to investigate whether P-selectin polymorphisms are associated with thrombosis in patients with antiphospholipid syndrome (APS). MATERIALS AND METHODS: The diagnosis and classification of APS were based on the report of an international workshop. Genomic DNA was extracted from citrated blood samples of all subjects. Three single nucleotide polymorphisms associated with the P-selectin coding region (S290N, c.1087G>A; N562D, c.1902G>A; T715P, c.2363A>C) were assessed. RESULTS: There were 26 APS (65%) patients with thrombosis. The number of patients without thrombosis was 14 (35%). The frequency of the N562D-DN genotype was significantly higher in patients with APS than in healthy controls (p=0.003). The frequency of this genotype was significantly higher in patients with APS with thrombosis compared with patients with no thrombosis (p=0.03). The N562D-NN genotype was found at a higher frequency in patients with APS than in healthy controls (p=0.004). CONCLUSION: Our results suggest that the N562D polymorphism of the DN genotype of P-selectin is associated with an increased risk of thrombosis in patients with APS.