Overall Approach of the laboratory The laboratory uses a translational research approach to study human malformations and overgrowth disorders. In the clinical arena (study HG200388-04), we operate several clinical research protocols to assess the range of severity, spectrum of malformations, natural history of pleiotropic developmental and overgrowth disorders, and therapeutic studies. We use the tools of modern molecular biology to determine the molecular pathogenesis of these disorders. These include high throughput sequencing, positional cloning, microarray expression and microarray CGH analysis, cell and tissue culture studies to assess cell biologic functions and abnormalities of gene products, and the creation and analysis of animal mouse models of human genetic disease, and now we have developed therapeutic protocols for several of these diseases. Overgrowth syndromes Building on our prior successes with PIK3CA-related fibroadipose overgrowth (Lindhurst et al, 2012) and Proteus syndrome (Lindhurst et al, 2011), we have now identified targets for therapeutic intervention. These discoveries have shown that the mutations that cause mosaic overgrowth also are major contributors to the pathogenesis of cancer. Because of the intense work in the pharmaceutical industry on cancer drugs, we have used two such agents for therapeutics of overgrowth. We are using single-cell cloned fibroblast lines from patients with Proteus syndrome to test therapeutic agents. By titrating dosage of agents against assays of cell death and proliferation we have measured the potential effect of these agents for future clinical development. These preclinical data supported our successful application for an IND to use this compound to treat affected patients. Modeling Proteus syndrome Following on our discovery of the cause of this disorder in 2011 we have been pursuing a strategy to model this disorder. While the Happle hypothesis (mosaicism for a mutation lethal in the non-mosaic state) is completely consistent with all recognized features of the disorder, it is impossible to prove this in human studies. To that end, we have undertaken efforts to create a mouse model of Proteus syndrome by creating a conditional knock-in allele for the p.Glu17Lys mutation (the mutation that affects all known patients with this disorder). We have made excellent progress on this project with extensive genetic engineering accomplished to make and validate the construct. The animals have been made and have been bred to cre-promoter constructs to answer our scientific questions. Genotype-Phenotype studies in mosaic disorders We have developed tissue sampling and culture methods coupled with custom-engineered mutation assays to detect the 5 known mutations in these genes in patients thought to be affected with this disorder and now routinely perform this as a patient screening method. This will allow us to support the activities in our clinical research project (HG200388-04) to reclassify these phenotypes. For any patients in whom this analysis is negative, we feed these samples into our next generation sequencing analysis pipeline using the intrapatient exome mosaic comparison approach that we have pioneered. Exome analysis of novel germline phenotypes In this past year we have used exome analysis to evaluate about 20 distinctive phenotypes, with the elucidation of probable genetic etiologies for 5 of these, which is quite similar to the success rates for other groups. We have determined the cause of a novel form of Noonan syndrome in the past year, a novel scarring/contraction syndrome, and a new cause of syndromic microphthalmia.

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14
Fiscal Year
2018
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Name
Human Genome Research
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Biesecker, Leslie G (2018) Mosaic disorders and the Taxonomy of Human Disease. Genet Med 20:800-801
Johnston, Jennifer J; van der Smagt, Jasper J; Rosenfeld, Jill A et al. (2018) Autosomal recessive Noonan syndrome associated with biallelic LZTR1 variants. Genet Med 20:1175-1185
Nathan, Neera R; Patel, Rachna; Crenshaw, Molly M et al. (2018) Pathogenetic insights from quantification of the cerebriform connective tissue nevus in Proteus syndrome. J Am Acad Dermatol 78:725-732
Biderman Waberski, Marta; Lindhurst, Marjorie; Keppler-Noreuil, Kim M et al. (2018) Urine cell-free DNA is a biomarker for nephroblastomatosis or Wilms tumor in PIK3CA-related overgrowth spectrum (PROS). Genet Med 20:1077-1081
Biesecker, Leslie G (2018) Response to Nakaguma et al. Genet Med :
Johnston, Jennifer J; Lee, Chanjae; Wentzensen, Ingrid M et al. (2017) Compound heterozygous alterations in intraflagellar transport protein CLUAP1 in a child with a novel Joubert and oral-facial-digital overlap syndrome. Cold Spring Harb Mol Case Stud 3:
Sapp, Julie C; Hu, Lian; Zhao, Jean et al. (2017) Quantifying survival in patients with Proteus syndrome. Genet Med 19:1376-1379
Yuan, Xuan; Li, Zhe; Baines, Andrea C et al. (2017) A hypomorphic PIGA gene mutation causes severe defects in neuron development and susceptibility to complement-mediated toxicity in a human iPSC model. PLoS One 12:e0174074
Nathan, Neera; Keppler-Noreuil, Kim M; Biesecker, Leslie G et al. (2017) Mosaic Disorders of the PI3K/PTEN/AKT/TSC/mTORC1 Signaling Pathway. Dermatol Clin 35:51-60
Wentzensen, Ingrid M; Johnston, Jennifer J; Patton, John H et al. (2016) Exome sequencing identifies a mutation in OFD1 in a male with Joubert syndrome, orofaciodigital spectrum anomalies and complex polydactyly. Hum Genome Var 3:15069

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