Angiomyolipoma Have Common Mutations in TSC2 but No Other Common Genetic Events

Renal angiomyolipoma are part of the PEComa family of neoplasms, and occur both in association with Tuberous Sclerosis Complex (TSC) and independent of that disorder. Previous studies on the molecular genetic alterations that occur in angiomyolipoma are very limited. We evaluated 9 angiomyolipoma for which frozen tissue was available from a consecutive surgical series. Seven of 8 samples subjected to RT-PCR-cDNA sequencing showed mutations in TSC2; none showed mutations in TSC1 or RHEB. Six of the seven mutations were deletions. We searched for 983 activating and inactivating mutations in 115 genes, and found none in these tumors. Similarly analysis for genomic regions of loss or gain, assessed by Affymetrix SNP6.0 analysis, showed no abnormalities. Loss of heterozygosity in the TSC2 region was commonly seen, except in patients with low frequency TSC2 mutations. We conclude that sporadic renal angiomyolipoma usually have mutations in TSC2, but not TSC1 or RHEB, and have no other common genomic events, among those we searched for. However, chromosomal translocations and gene fusion events were not assessed here. TSC2 inactivation by mutation is a consistent and likely necessary genetic event in the pathogenesis of most angiomyolipoma

Real-Time Monitoring of Tumorigenesis, Dissemination, & Drug Response in a Preclinical Model of Lymphangioleiomyomatosis/Tuberous Sclerosis Complex

Background: TSC2-deficient cells can proliferate in the lungs, kidneys, and other organs causing devastating progressive multisystem disorders such as lymphangioleiomyomatosis (LAM) and tuberous sclerosis complex (TSC). Preclinical models utilizing LAM patient-derived cells have been difficult to establish. We developed a novel animal model system to study the molecular mechanisms of TSC/LAM pathogenesis and tumorigenesis and provide a platform for drug testing. Methods and Findings: TSC2-deficient human cells, derived from the angiomyolipoma of a LAM patient, were engineered to co-express both sodium-iodide symporter (NIS) and green fluorescent protein (GFP). Cells were inoculated intraparenchymally, intravenously, or intratracheally into athymic NCr nu/nu mice and cells were tracked and quantified using single photon emission computed tomography (SPECT) and computed tomography (CT). Surprisingly, TSC2-deficient cells administered intratracheally resulted in rapid dissemination to lymph node basins throughout the body, and histopathological changes in the lung consistent with LAM. Estrogen was found to be permissive for tumor growth and dissemination. Rapamycin inhibited tumor growth, but tumors regrew after the drug treatment was withdrawn. Conclusions: We generated homogeneous NIS/GFP co-expressing TSC2-deficient, patient-derived cells that can proliferate and migrate in vivo after intratracheal instillation. Although the animal model we describe has some limitations, we demonstrate that systemic tumors formed from TSC2-deficient cells can be monitored and quantified noninvasively over time using SPECT/CT, thus providing a much needed model system for in vivo drug testing and mechanistic studies of TSC2-deficient cells and their related clinical syndromes

Multicenter Phase 2 Trial of Sirolimus for Tuberous Sclerosis: Kidney Angiomyolipomas and Other Tumors Regress and VEGF- D Levels Decrease

Tuberous sclerosis (TSC) related tumors are characterized by constitutively activated mTOR signaling due to mutations in TSC1 or TSC2.We completed a phase 2 multicenter trial to evaluate the efficacy and tolerability of the mTOR inhibitor, sirolimus, for the treatment of kidney angiomyolipomas.36 adults with TSC or TSC/LAM were enrolled and started on daily sirolimus. The overall response rate was 44.4% (95% confidence intervals [CI] 28 to 61); 16/36 had a partial response. The remainder had stable disease (47.2%, 17/36), or were unevaluable (8.3%, 3/36). The mean decrease in kidney tumor size (sum of the longest diameters [sum LD]) was 29.9% (95% CI, 22 to 37; n = 28 at week 52). Drug related grade 1-2 toxicities that occurred with a frequency of >20% included: stomatitis, hypertriglyceridemia, hypercholesterolemia, bone marrow suppression (anemia, mild neutropenia, leucopenia), proteinuria, and joint pain. There were three drug related grade 3 events: lymphopenia, headache, weight gain. Kidney angiomyolipomas regrew when sirolimus was discontinued but responses tended to persist if treatment was continued after week 52. We observed regression of brain tumors (SEGAs) in 7/11 cases (26% mean decrease in diameter), regression of liver angiomyolipomas in 4/5 cases (32.1% mean decrease in longest diameter), subjective improvement in facial angiofibromas in 57%, and stable lung function in women with TSC/LAM (n = 15). A correlative biomarker study showed that serum VEGF-D levels are elevated at baseline, decrease with sirolimus treatment, and correlate with kidney angiomyolipoma size (Spearman correlation coefficient 0.54, p = 0.001, at baseline).Sirolimus treatment for 52 weeks induced regression of kidney angiomyolipomas, SEGAs, and liver angiomyolipomas. Serum VEGF-D may be a useful biomarker for monitoring kidney angiomyolipoma size. Future studies are needed to determine benefits and risks of longer duration treatment in adults and children with TSC.Clinicaltrials.gov NCT00126672

ANIMAL MODELS FOR THE STUDY OF LEISHMANIASIS IMMUNOLOGY

Leishmaniasis remains a major public health problem worldwide and is classified as Category I by the TDR/WHO, mainly due to the absence of control. Many experimental models like rodents, dogs and monkeys have been developed, each with specific features, in order to characterize the immune response to Leishmania species, but none reproduces the pathology observed in human disease. Conflicting data may arise in part because different parasite strains or species are being examined, different tissue targets (mice footpad, ear, or base of tail) are being infected, and different numbers (“low” 1×102 and “high” 1×106) of metacyclic promastigotes have been inoculated. Recently, new approaches have been proposed to provide more meaningful data regarding the host response and pathogenesis that parallels human disease. The use of sand fly saliva and low numbers of parasites in experimental infections has led to mimic natural transmission and find new molecules and immune mechanisms which should be considered when designing vaccines and control strategies. Moreover, the use of wild rodents as experimental models has been proposed as a good alternative for studying the host-pathogen relationships and for testing candidate vaccines. To date, using natural reservoirs to study Leishmania infection has been challenging because immunologic reagents for use in wild rodents are lacking. This review discusses the principal immunological findings against Leishmania infection in different animal models highlighting the importance of using experimental conditions similar to natural transmission and reservoir species as experimental models to study the immunopathology of the disease

Mutational analysis of the tuberous sclerosis gene TSC2 in patients with pulmonary lymphangioleiomyomatosis

Pulmonary lymphangioleiomyomatosis (LAM) is a rare disorder limited almost exclusively to women of reproductive age. LAM affects about 5% of women with tuberous sclerosis complex (TSC). LAM also occurs in women who do not have TSC (sporadic LAM). TSC is a tumour suppressor gene syndrome characterised by seizures, mental retardation, and tumours in the brain, heart, and kidney. Angiomyolipomas, which are benign tumours with smooth muscle, fat, and dysplastic vascular components, are the most common renal tumour in TSC. Renal angiomyolipomas also occur in 63% of sporadic LAM patients. We recently found that 54% of these angiomyolipomas have TSC2 loss of heterozygosity, leading to the hypothesis that sporadic LAM is genetically related to TSC. In this study, we screened DNA from 21 women with sporadic LAM for mutations in all 41 exons of TSC2. Twelve of the patients had known renal angiomyolipomas. No TSC2 mutations were detected. We did find three silent TSC2 polymorphisms. We conclude that patients with sporadic LAM, including those with renal angiomyolipomas, do not have a high frequency of germline mutations in the coding region of TSC2.


Keywords: TSC2; pulmonary lymphangioleiomyomatosi

Cotton Rat (Sigmodon hispidus) Signaling Lymphocyte Activation Molecule (CD150) Is an Entry Receptor for Measles Virus

Cotton rats (Sigmodon hispidus) replicate measles virus (MV) after intranasal infection in the respiratory tract and lymphoid tissue. We have cloned the cotton rat signaling lymphocytic activation molecule (CD150, SLAM) in order to investigate its role as a potential receptor for MV. Cotton rat CD150 displays 58% and 78% amino acid homology with human and mouse CD150, respectively. By staining with a newly generated cotton rat CD150 specific monoclonal antibody expression of CD150 was confirmed in cotton rat lymphoid cells and in tissues with a pattern of expression similar to mouse and humans. Previously, binding of MV hemagglutinin has been shown to be dependent on amino acids 60, 61 and 63 in the V region of CD150. The human molecule contains isoleucine, histidine and valine at these positions and binds to MV-H whereas the mouse molecule contains valine, arginine and leucine and does not function as a receptor for MV. In the cotton rat molecule, amino acids 61 and 63 are identical with the mouse molecule and amino acid 60 with the human molecule. After transfection with cotton rat CD150 HEK 293 T cells became susceptible to infection with single cycle VSV pseudotype virus expressing wild type MV glycoproteins and with a MV wildtype virus. After infection, cells expressing cotton rat CD150 replicated virus to lower levels than cells expressing the human molecule and formed smaller plaques. These data might explain why the cotton rat is a semipermissive model for measles virus infection