A pilot study of next generation sequencing-liquid biopsy on cell-free DNA as a novel non-invasive diagnostic tool for Klippel-Trenaunay syndrome.

OBJECTIVES: Somatic mosaicism of PIK3CA gene is currently recognized as the molecular driver of Klippel-Trenaunay syndrome. However, given the limitation of the current technologies, PIK3CA somatic mutations are detected only in a limited proportion of Klippel-Trenaunay syndrome cases and tissue biopsy remains an invasive high risky, sometimes life-threatening, diagnostic procedure. Next generation sequencing liquid biopsy using cell-free DNA has emerged as an innovative non-invasive approach for early detection and monitoring of cancer. This approach, overcoming the space-time profile constraint of tissue biopsies, opens a new scenario also for others diseases caused by somatic mutations. METHODS: In the present study, we performed a comprehensive analysis of seven patients (four females and three males) with Klippel-Trenaunay syndrome. Blood samples from both peripheral and efferent vein from malformation were collected and cell-free DNA was extracted from plasma. Tissue biopsies from vascular lesions were also collected when available. Cell-free DNA libraries were performed using Oncomine™ Pan-Cancer Cell-Free Assay. Ion Proton for sequencing and Ion Reporter Software for analysis were used (Life Technologies, Carlsbad, CA, USA). RESULTS: Cell-free circulating DNA analysis revealed pathogenic mutations in PIK3CA gene in all patients. The mutational load was higher in plasma obtained from the efferent vein at lesional site (0.81%) than in the peripheral vein (0.64%) leading to conclude for a causative role of the identified variants. Tissue analysis, available for one amputated patient, confirmed the presence of the mutation at the malformation site at a high molecular frequency (14-25%), confirming its causative role. CONCLUSIONS: Our data prove for the first time that the cell-free DNA-next generation sequencing-liquid biopsy, which is currently used exclusively in an oncologic setting, is indeed the most effective tool for Klippel-Trenaunay syndrome diagnosis and tailored personalized treatment.

High levels of serum sclerostin and DKK1 in a case of Klippel-Trénaunay syndrome.

Klippel-Trénaunay syndrome (KTS) is described as a complex syndrome characterized by various combinations of capillary, venous, and lymphatic malformations associated with bone and soft tissue hypertrophy. We report a case of a 67-year-old postmenopausal Caucasian women with KTS that shows elevated levels of sclerostin and Dickkopf-related protein 1 (DKK1). Dual-energy X-ray absorptiometry (DXA) BMD T-scores at lumbar spine and femur were normal. Serum calcium and phosphorus levels were consistently normal, 25-hydroxyvitamin D (25OHD) < 30 ng/mL, and normal parathyroid hormone (PTH). Turnover markers (serum osteocalcin [OCN], and carboxy-terminal cross-linking telopeptide of type 1 collagen [CTx]) were in the reference limits. It is interesting to note that the serum levels of sclerostin and DKK-1 were significantly higher in our patient with KTS than in a healthy volunteer (control), without impact on bone mineral density and bone formation markers. In fact, in our patient, the BMD at lumbar spine and femur was normal, and osteocalcin was not suppressed. Based on what is known, we would have expected to find low levels of the inhibitors of the Wnt system, perhaps we can explain the data as a response to the compensation for ß-catenin hyper-transformation.

[Klippel-Trenaunay syndrome associated with antithrombin III deficiency]. / Syndrome de Klippel-Trenaunay et déficit en antithrombine III A propos d'une observation.

Klippel-Trenaunay syndrome (KTS) is a rare phakomatosis characterized by cutaneous hemangiomata, venous varicosities and bone and soft tissue hypertrophy also of the affected limb. Central nervous system involvement is rare, arising from a malformation or from coagulation disorders. We report the case of a patient presenting a KTS with stroke. The biological assessment revealed antithrombin III deficiency. Although rare, antithrombin III deficiency should be kept in mind in KTS patients with neurological involvement.

[Autologous transfusion of UV-irradiated blood in the complex treatment of children with dysplasia of deep veins of the extremities]. / Autotransfuziia obluchennoi ul'travioletom krovi v kompleksnom lechenii detei s displaziei glubokikh ven konechnostei.

Under examination there were 12 patients aged from 5 till 14 years. An investigation of hemostatic potential of blood in the diseased extremity and peripheral blood was performed. A mosaic character of blood coagulative alterations in this category of patients was detected. Autotransfusion of UV-irradiated blood is a pathogenetically grounded and accessible method of correction of hemocoagulation and may be included in the complex treatment of patients with dysplasia of the profound veins of lower extremities.

Klippel-Trénaunay syndrome (KTS) evaluated with bone and labeled RBC scintigraphy.

This is a case of a 4-year-old female child with gross left lower extremity deformity detected at birth, including a giant cutaneous hemangioma from flank to foot. She was found to have no other associated abnormalities and was diagnosed with Klippel-Trénaunay syndrome. This is a rare entity and literature regarding use of radionuclide imaging techniques to evaluate this is scarce. We present here bone and red cell scans performed to evaluate whether there was osseous involvement or only soft tissue disease, which showed classic signs of this condition. Patient photographs also show the syndrome's typical appearance.

Venous thromboembolism and prothrombotic parameters in Klippel-Trenaunay syndrome.

BACKGROUND: In Klippel-Trenaunay syndrome (KTS), a congenital combined vascular (capillary, venous and lymphatic) malformation with localised disturbed growth, venous thromboembolisms (VTEs) are frequently reported in small cohorts. DESIGN AND METHODS: We quantified the frequency of VTE by screening a large KTS-patient cohort with duplex compression ultrasonography. Additionally, we performed a case-control study to evaluate whether coagulation alterations were related to VTE and magnitude of vascular malformations as quantified by magnetic resonance imaging (MRI). RESULTS: Twenty-nine (39%) of 75 patients had signs of current or previous VTE, including superficial venous thrombosis, six (8%) of whom had a deep venous thrombosis or a pulmonary embolism. Compared with 105 controls, 54 adult patients (both: median age 33 years) had higher plasma levels of D-dimer, medians 266 (IQR 195-366) versus 457 (IQR 270-3840) mg÷l (p.

Elevated D-dimer level in the differential diagnosis of venous malformations.

OBJECTIVE: To evaluate if elevated D-dimer level is specific for venous malformations (VMs) and thus useful for differential diagnosis, which can be problematic even in specialized interdisciplinary centers. Localized intravascular coagulopathy, characterized by elevated D-dimer levels, has been observed in approximately 40% of patients with VMs. DESIGN: Prospective convenience sample accrued from 2 interdisciplinary sites. SETTING: Two interdisciplinary centers for vascular anomalies in Brussels, Belgium, and Caen, France PARTICIPANTS: The study population comprised 280 patients with clinical data, Doppler ultrasonograms (for 251 patients), and coagulation parameter measurements. Main Outcome Measure Measurement of D-dimer levels. RESULTS: A VM was diagnosed in 195 of 280 patients (69.6%), and 83 of them had elevated D-dimer levels; the sensitivity of D-dimer dosage was 42.6% (95% confidence interval, 35.6%-49.5%). Among the 85 patients without VM, D-dimer levels were elevated only in 3 patients; the specificity of the dosage was 96.5% (95% confidence interval, 92.5%-100%). CONCLUSIONS: Elevated D-dimer level is highly specific for VMs (pure, combined, or syndromic), and therefore this easy and inexpensive biomarker test should become part of the clinical evaluation of vascular anomalies. It can detect hidden VMs and help differentiate glomuvenous malformation (normal D-dimer levels) from other multifocal venous lesions. Elevated D-dimer level also differentiates a VM from a lymphatic malformation. Moreover, slow-flow Klippel-Trenaunay syndrome (capillaro-lymphatico-venous malformation with limb hypertrophy) can be distinguished from fast-flow Parkes Weber syndrome (capillary malformation with underlying multiple microfistulas and limb hypertrophy). For these reasons, D-dimer level measurement is a useful complementary tool for diagnosing vascular anomalies in everyday practice.

The Klippel Trenaunay Weber Syndrome presenting with cutaneous bleeding.

A patient with the Klippel Trenaunay Weber syndrome presented with prolonged bleeding from the finger tips during strenuous exercise. Tissue fibrinolysis studies showed increased fibrinolytic activity suggesting that the blood vessels in the affected limb may be functionally as well as anatomically abnormal.

Severe hemorrhage complicating the Klippel-Trénaunay-Weber syndrome.

The Klippel-Trénaunay-Weber (KTW) syndrome is a congenital disorder of angiogenesis characterized by macular nevus, skeletal and soft tissue hypertrophy, venous varicosities, and arteriovenous fistulas. Disseminated intravascular coagulation (DIC) and the Kasabach-Merritt syndrome, a consumptive coagulopathy with thrombocytopenia, are both associated with the KTW syndrome. We describe a 30-year-old woman with KTW syndrome and Kasabach-Merritt syndrome who had DIC with severe hemorrhage after a routine gynecologic procedure. The bleeding was controlled with the use of intravenous low-dose heparin and antithrombin III.

[Thromboembolic disease and congenital venous abnormalities]. / Maladie thrombo-embolique et anomalies veineuses congénitales.

A pulmonary embolus and a bilateral thrombosis of the internal carotid artery caused the sudden death of two young adults with a congenital venous abnormality. The study of 49 patients suffering from a Klippel-Trenaunay syndrome showed that 11 out of 49 patients, or 22.5 p. cent had thrombosis problems: namely 7 pulmonary emboli and 8 deep thrombophlebitis. Phlebography did not permit to discover any specific anatomical abnormality. 5 out of 11 patients presented avalvular and dilated deep veins. 6 out of 11 presented angioma of the pelvis. In the group of patients with Klippel-Trenaunay who never presented a deep venous thrombosis, the same signs are noted in one third of the cases. It is possible that coagulation insufficiency may explain the high number of thrombophlebitis. However, an extended analysis of 11 patients did not permit to discover any abnormality. But, subsequently, a study of the fibrinolytic activity demonstrated a normal fibrinogen level, while the fibrinopeptide A level was markedly elevated in each case, with an abnormal thrombin activity.