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Wo 2008/022335 A2 (12) INTERNATIONAL APPLICATION PUBLISHED UNDER THE PATENT COOPERATION TREATY (PCT) (19) World Intellectual Property Organization International Bureau (43) International Publication Date (10) International Publication Number 21 February 2008 (21.02.2008) PCT WO 2008/022335 A2 (51) International Patent Classification: Pemburton Avenue, #806, Toronto, Ontario M2M 4K8 C12Q 1/68 (2006.01) (CA). (21) International Application Number: (74) Agents: MYERS, Louis et al.; Fish & Richardson P.C., PCT/US2007/076248 P.O. Box 1022, Minneapolis, Minnesota 55440-1022 (US). (22) International Filing Date: 17 August 2007 (17.08.2007) (81) Designated States (unless otherwise indicated, for every (25) Filing Language: English kind of national protection available): AE, AG, AL, AM, AT,AU, AZ, BA, BB, BG, BH, BR, BW, BY, BZ, CA, CH, (26) Publication Language: English CN, CO, CR, CU, CZ, DE, DK, DM, DO, DZ, EC, EE, EG, (30) Priority Data: ES, FT, GB, GD, GE, GH, GM, GT, HN, HR, HU, ID, IL, 60/838,662 18 August 2006 (18.08.2006) US IN, IS, JP, KE, KG, KM, KN, KP, KR, KZ, LA, LC, LK, 60/845,564 19 September 2006 (19.09.2006) US LR, LS, LT, LU, LY, MA, MD, ME, MG, MK, MN, MW, MX, MY, MZ, NA, NG, NI, NO, NZ, OM, PG, PH, PL, (71) Applicants (for all designated States except US): THE PT, RO, RS, RU, SC, SD, SE, SG, SK, SL, SM, SV, SY, BRIGHAM AND WOMEN'S HOSPITAL, INC. TJ, TM, TN, TR, TT, TZ, UA, UG, US, UZ, VC, VN, ZA, [US/US]; 75 Francis Street, Boston, Massachusetts 021 15 ZM, ZW (US). BETH ISRAEL DEACONESS MEDICAL CEN¬ TER, INC. [US/US]; 330 Brookline Avenue, Boston, (84) Designated States (unless otherwise indicated, for every Massachusetts 02215 (US). kind of regional protection available): ARIPO (BW, GH, GM, KE, LS, MW, MZ, NA, SD, SL, SZ, TZ, UG, ZM, (72) Inventors; and ZW), Eurasian (AM, AZ, BY, KG, KZ, MD, RU, TJ, TM), (75) Inventors/Applicants (for US only): NEEL, Benjamin European (AT,BE, BG, CH, CY, CZ, DE, DK, EE, ES, FI, G. [US/CA]; 48 Glen Road, Toronto, Ontario M4W2V1 FR, GB, GR, HU, IE, IS, IT, LT,LU, LV,MC, MT, NL, PL, (CA). ROBERTS, Amy E. [US/US]; 12 Allston Street, PT, RO, SE, SI, SK, TR), OAPI (BF, BJ, CF, CG, CI, CM, #1, Charlestown, Massachusetts 02129 (US). KUCHER- GA, GN, GQ, GW, ML, MR, NE, SN, TD, TG). LAPATI, Raju [US/US]; 6 Wildflower Lane, Weston, Massachusetts 02493 (US). SWANSON, Kenneth D. Published: [US/US]; 1454 Beacon St., Unit 441, Brookline, Mass — without international search report and to be republished achusetts 02446 (US). ARAKI, Toshiyuki [JP/CA]; 8 upon receipt of that report (54) Title: DIAGNOSIS AND TREATMENT OF NOONAN SYNDROME AND NEOPLASTIC DISORDERS (57) Abstract: Methods and compositions for diagnosing and treating Noonan syndrome and neoplastic disorders are provided herein. DIAGNOSIS AND TREATMENT OF NOONAN SYNDROME AND NEOPLASTIC DIS ORDERS CROSS-REFERENCE TO RELATED AFF[JCAHONS This application claims the benefit of priority of O .S.S.N. 60/838,662, filed August 1 , 2006, ami U.S.S.N. 60/845,564, filed September 19, 2006. The coalents of the prior applications are hereby incorporated by reference in their entirety. FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT The work described herein was funded, in part through grants from the National Institutes oi' Health (grants R37CA491 52, DE i 6140, and MO !-RR02 172) The United States government may, therefore, have certain rights in the invention TECHNICAL FIELD This invention relates to methods and compositions for diagnosis and t at e t of genetic disorders, miά more particularly to diagnosis and treatment of Noonan syndrome, and neoplastic disorders. BACKGROUND Noonan syndrome (NS) is the most comm υn single-gene cause of congenital heart disease (CHD), and also frequently includes short stature, characteristic facial features, learning problems, and an increased risk of certain lcukemias (Tartaglia d Oeib, Anna Rev Genomics Hum Genet, 6:45-68, 2005). Consistent with its autosomal dominant inheritance pattern, gain-of- function mutations in the PTPNIl gene, encoding the tyrosine phosphatase SHP2, cause -50% of NS cases. SHP2 is required for fall activation of the RAS/HRK. MAP kinase (MAPK) cascade downstream of most growth factor and cytokine receptors, and NS mutants enhance ERK activation ex vivo and in ic (Fragale et al.. Num. MuL, 23: 267- 277, 2004; Kontaridis et al., Biol Chem., 281:6785-6792, 2005; Araki et aL Nat Med., 10:849-857, 2004). ATi-ISmutations account for <5% of NS (Scfaubberi ct aL Ηat. Genet., 38:331-336, 2006), but the gεnε(s) responsible for the remainder of NS cases remain unknown. SUMMARY The invention is based, i part, on the discovery of mutations within the Iranian So oCsevcπlcss 1 (SOS i) gene which are associated with hu disease. The invention provides, inter alia, methods and compositions for diagnosing and treating human disorders including NS ! neoplastic disorders In o aspect, the invention features a method for diagnosing i a subject, or identifying a subject at risk for, Noonan syndrome (NS). The et od includes, for example, determining if one or o e imitations are present m a SOS gene (e.g., SOSl) of the subject, h rei the presence of one or more .mutations indicates that the subject is affected with, or at risk for, NS. I one embodiment, \\\Q subject is subject who presents with one or more phenoiyp c characteristics of NS. Phsnotypic characteristics of NS include dysmorphic facial features (e.g., broad forehead, hypertelorism down-slanting palpebral fissures, highly arched palate, low set mid posteriorly rotated cars), proportionate short stature, pectus deformity, cryptorchidism, developmental delay genitourinary malformations, bleeding disorders, lymphatic dysplasia, growth failure, and cardiac defects (e.g., hypertrophic cardiomyopathy, pulmonic stenosis, atrial septal defect, and aortic coarctation). In one embodiment, the subject has been screened for a mutation i the PTFN 11 gene (e.g., the subject has been identified as lacking a mutation in the PTFNH gene, prior to screening for SOSl mutations). The method can further include determining whether a PTP 11 and/or a KRAS gene of the subj ect has a mutation. For example, the method includes screening for mutations in SOS! genes, PTPN 11, and KRAS genes of the subject. in various embodiments of the methods, subjects are evaluated for mutations which are substitutions, deletions, or insertions of one or more nucleotides in a SOS 1 gene. In o embodiment, the mutation includes a mutation of single nucleotide, e.g., a substitution, deletion, or insertion of a single nucleotide. In o e embodiment, the mutation includes a missense mutation. Subjects may be evaluated for mutations which are chromosomal rearrangements (e.g., translocations or deletions, ch as translocations or deletions which activate SOS 1 by juxtaposing a regulatory sequence with a SOS 1 coding sequence). Exemplary mutations indicative of NS (or risk for NS) include mutations at o e of the following nucleotide positions of the SOS 1 sequence of SEQ ID NO: 1; 797, 806, 925. !OiO, 1358, 642, 1654, 1964, and 253(S. In some embodiments, the mmation is other tbars one of the following imitations: n insertion between nucleotides 3248 and 3249, or a mutation at nucleotide 3032. Io one embodiment, the mutation indicative of NS s a substitution in the SOSI coding sequence, e.g., corresponding to one of the following substitutions in SEQ ID Oύ : 797OA, 8061X3, 925OT, 1010A>G, 1358G>C, 1642A>C, 1654A>G, 1964OX or 2536G>A. In one embodiment, the mutation in the SOSl gene results in ihe substitution, deletion or insertion of one or more amino acids of Ihe polypeptide encoded by the gene. For example, the mutation results in a mutation at one or more of the following amnio acid positions in the SOS I polypeptide of SEQ B NG:2: 7766, M269, D309, Y337, G434, S548, R552, P655, or E846. The substitution can include one of the following substitutions: T266K, M269R, D309Y, Y337C, G434R, S548R, R552G, P655L, or E846 In one embodiment, the mutation Ui the SOS 1 gene results in a mutation in one of the following domains of the polypeptide encoded by the g e: the Db! Homology (DH) domain, the Pkekstrin Homology (PH) domain, the Helical Linker (HL) domain, fee Ras Exchange MoUf(REM) domain, or the Cdc25 domain. In one embodiment, the mutation in the SOSl gene results in a increased level of expression or activity f the polypeptide encoded by the gene. For example, ihe polypeptide encoded by the gene mediates enhanced Ras, Erk, and/or R&c activation, relative to a control (e.g., relative to a wild type SOS 1polypeptide). hi one embodiment, the mutation includes a mutation in an exon of the SC)SI gene (e.g. ., a mutation i exon 6, exon 7, exon 8, exon 9, exon 10, exon !2 exon 16, or exon 19}. In another embodiment the mutation is a mutation in a promoter, enhancer, untranslated region (UTR), or intron of the gene (e.g., a 3 UTR 5TiTR). The e od of determining whether &subject has a mutation i a SOSl gene can include determining he identity of at least o e nucleotide in the SOSl gene of the subject (e.g., wherein the nucleotide in the SOSl gene is m an exon, intron, regulatory, 3'UTR., and/or 5'UTR region of the gene, e.g., wherein a the identify of.it least 50, 100, 250, 500, 1000, 2500, or 50(50 nucleotides of the SOSI gene is determined). Alternatively, or in addition, the method can include determining whether the subject contains a marker (e.g., tx polymorphism) which i linked to a SOSi mutation.
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