US 2011 0312520A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2011/0312520 A1 Kennedy et al. (43) Pub. Date: Dec. 22, 2011 (54) METHODS AND COMPOSITIONS FOR Publication Classification DAGNOSING CONDITIONS (51) Int. Cl. (75) Inventors: Giulia C. Kennedy, San Francisco, CI2O I/68 (2006.01) CA (US); Darya I. Chudova, San C40B 30/04 (2006.01) Jose, CA (US); Jonathan I. Wilde, C40B 30/00 (2006.01) Burlingame, CA (US); James G. Veitch, Berkeley, CA (US); Bonnie H. Anderson, Half Moon Bay, CA (US) (52) U.S. Cl. ................ 506/9:435/6.14; 435/6. 12:506/7 (73) Assignee: Veracyte, Inc., South San Francisco, CA (US) (21) Appl. No.: 13/105,756 (57) ABSTRACT (22) Filed: May 11, 2011 The present invention relates to compositions, kits, and meth Related U.S. Application Data ods for molecular profiling for diagnosing disease conditions. .S. App In particular, the present invention provides molecular pro (60) Provisional application No. 61/333,717, filed on May files associated with thyroid cancer and other cancers, meth 11, 2010, provisional application No. 61/389,810, ods of relating molecular profiles to a diagnosis, and related filed on Oct. 5, 2010. compositions. Patent Application Publication Dec. 22, 2011 Sheet 1 of 28 US 2011/0312520 A1 FIGURE 1A Expression Analysis Gene Expression Level(s) Training/Reference Samples Biomarker Set/ Classifier 1 no match Training/Reference Samples Biomarker Set/ Classifier 2 Patent Application Publication Dec. 22, 2011 Sheet 2 of 28 US 2011/0312520 A1 &&&&#3:********** &&&&&&&&&&& Patent Application Publication Dec. 22, 2011 Sheet 3 of 28 US 2011/0312520 A1 FIGURE 1C 201 Module 1 (Module1.pm) Paralelized APT processes checksum apt-Cel-transformer e apt-probeset-Summarize (ps) apt-probeset-summarize (mps) apt-tSV-join Invocation interface Directory locking (LockDirpm) ConfigFile Handler (ConfigFile.pm), messages File Process Execution Patent Application Publication Dec. 22, 2011 Sheet 4 of 28 US 2011/0312520 A1 FIGURE 2 Gene Biomarker Panels for Diagnosing a Thyroid Condition Normal Thyroid (NML) Lymphocytic, Autoimmune Thyroiditis (LCT) Nodular Hyperplasia (NHP) Follicular Thyroid Adenoma (FA) Hurthle Cell Thyroid Adenoma (HA) Parathyroid (non thyroid tissue) (PTA) Anaplastic Thyroid Carcinoma (ATC) Follicular Thyroid Carcinoma (FC) Hurthle Cell Thyroid Carcinoma (HC) Papillary Thyroid Carcinoma (PTC) Follicular Variant of Papillary Carcinoma (FVPTC) Medullary Thyroid Carcinoma (MTC) Renal Carcinoma metastasis to the Thyroid (RCC) Melanoma metastasis to the Thyroid (MMN) B cell Lymphoma metastasis to the Thyroid (BCL) Breast Carcinoma metastasis to the Thyroid (BCA) Patent Application Publication Dec. 22, 2011 Sheet 5 of 28 US 2011/0312520 A1 FIGURE 3 Optimized Gene Classification Panels of Diagnostic Utility in Thyroid FNA Classification Subtype-specific Classifier Number of Markers Per Panel Panel Components MTC RCC PTA BCA MMN HA & HC 33 Benign/Suspicious 42 denotes the combination of two panels -HA and HC- into a single panel. denotes the combination of eight panels -FA, FC, NHP, PTC, FVPTC, LCT, HA, HC- into a single panel. Patent Application Publication Dec. 22, 2011 Sheet 6 of 28 US 2011/0312520 A1 FIGURE 4A TC) GENE Gene Description Classification Paned 336427 CALCA calcitonin-related polypeptide alpha MTC carcinoembryonic antigen-related cell adhesion 383434 CEACAMS molecule 5 MTC 35.94003 SCO3 secretogramin III - M - 2S854.00 SCN2A sodium channel, voltage-gated, type II, alpha subunit MTC sodium channel, voltage-gated, type IX, alpha 25854OO SCN9A subunit MTC 380S64 SYT4 synaptotagmin IV MTC 2923928 FABP7 fatty acid binding protein 7, brain RCC 3393446 FXYD2 FXYD domain containing ion transport regulator 2 RCC 288337 HAWCR hepatitis. A virus cellular receptor RCC 288337 LOCOOO1266 hepatitis. A virus cellular receptor pseudogene RCC 3428,225 NR14 nuclear receptor subfamily l, group II, member 4 RCC 2479698 PREPL prolyl endopeptidase-like RCC Solute carrier family 3 (cystine, dibasic and neutral amino acid transporters, activator of cystine, dibasic 2479698 SLC3A1 and neutral amino acid transport), member 1 RCC 3.1597.54 DMRT2 doublesex and mab-3 related transcription factor 2 PTA 2941690 GCM2 glial cells missing homolog 2 (Drosophila) PTA 33636.86 KDINS220 kinase D-interacting Substrate, 220kDa PTA 3484.895 KL klotho PTA 3363.686 PH parathyroid hormone PTA 2894790 SYCPL synaptonemal complex protein 2-like PTA 3O39830 AGR3 anterior gradient homolog 3 (Xenopus laevis) BCA 3264997 C10orfs chromosome 10 open reading frame 81 BCA V-myb myeloblastosis viral oncogene homolog 29268O2 MYB (avian) BCA 391.2079 SYCP2 synaptonema complex protein 2 BCA 243.63 WTCN1 V-set domain containing T cell activation inhibitor 1 BCA 3811949 CDII19 cadherin l9, type 2 MMN Patent Application Publication Dec. 22, 2011 Sheet 7 of 28 US 2011/0312520 A1 FIGURE 4B TCD GENE Gene Description Classification Panel 3.1626 MLANA melan-A MMN 39.354.86 SOOB S100 calcium binding protein B MMN 3457336 SLV silver homolog (mouse) MMN 33438.32 TYR tyrosinase (oculocutaneous albinism IA) MMN 3343832 TYRL tyrosinase-like (pseudogene) MMN HA & HC, 256.6848 AFF3 AF4/FMR2 family, member 3 Benign.Suspicious aminoacyl tRNA synthetase complex-interacting HA & HC. 2988882 AIMP2 multifunctional protein 2 Benign/Suspicious HA & HC, 316.933 ALDHB aldehyde dehydrogenasc 1 family, member Bl Bcnign.Suspicious HA & HC, 2.98466 BRP44L brain protein 44-like Benign/Suspicious HA & HC. 28224.92 C5orf30 chromosome 5 open reading frame 30 Benign/Suspicious HA & HC, 3326635 CD44 CD44 molecule (Indian blood group) Benign.Suspicious HA & HC, 2750627 CPE carboxypeptidase E Benign.Suspicious IIA & IIC, 3O4200 CYCS cytochrome c, Somatic Benign/Suspicious HA & HC, 3122678 DEFB1 defensin, beta 1 Benign.Suspicious HA & HC, 27393O8 EGF epidermal growth factor (beta-urogastrone) Benign.Suspicious eukaryotic translation initiation factor 2-alpha kinase HA & HC. 2988882 EIF2AK 1 Benign/Suspicious HA & HC, 36O3932 FAH fumarylacetoacetate hydrolase (fumarylacetoacetase) Benign.Suspicious HA & HC, 2970897 FRK fyn-related kinase Benign.Suspicious HA & HC. 3212008 FRMD3 FERM domain containing 3 Benign/Suspicious glutamic-oxaloacetic transaminasc 1, Soluble HA & HC, 33O2990 GOT (aspartate aminotransferase 1) Benign.Suspicious hydroxysteroid (17-beta) dehydrogenase 6 homolog HA & HC, 341 7703 ISD 736 (mouse) Benign/Suspicious HA & HC. 28775.08 HSPA9 heat shock 70kDa protein 9 (mortalin) Benign/Suspicious Patent Application Publication Dec. 22, 2011 Sheet 8 of 28 US 2011/0312520 A1 FIGURE 4C TC) GENE Gene Description Classification Panel HA & HC, 27O8922 GF2BP2 insulin-like growth factor 2 mRNA binding protein 2 Bcnign/Suspicious HA & HC, 2604998 IQCA IQ motif containing with AAA domain Benign.Suspicious integrin, beta 3 (platelet glycoprotein IIIa, antigen HA & HC, 3724.545 TGB3 CD61) Benign/Suspicious potassium inwardly-Tecifying channel, subfamily J, HA & HC, 3397.774 KCNJ member 1 Bcnign/Suspicious HA & HC, 2604998 LOCOO29258 hypothetical protein LOC10029258 Benign.Suspicious IIA & IIC, 3OO9299 MDI2 malate dehydrogenase 2, NAD (mitochondrial) Benign.Suspicious HA & HC, 3654699 NUPR1 nuclear protein, transcriptional regulator, 1. Benign/Suspicious HA & HC, 4O2O655 ODZ. odz, odd Oz ten-m homolog l(Drosophila) Benign/Suspicious HA & HC, 3970833 PDHA pyruvate dehydrogenase (lipoamide) alpha 1 Benign.Suspicious 6-phosphofructo-2-kinase/fructose-2,6- HA & HC, 237 7094 PFKFB2 biphosphatase 2 Benign/Suspicious HA & HC, 3278198 PHYH phytanoyl-CoA 2-hydroxylase Benign/Suspicious protein phosphatase 2 (formerly 2A), regulatory HA & HC, 288OO51 PPP2R2B Subunit B, beta isoform Benign.Suspicious HA & HC, 39598.62 PWALB parvalbumin Bcnign/Suspicious poliovirus receptor-related 2 (herpesvirus entry HA & HC, 2688499 PVRL2 mediator B) Benign.Suspicious IIA & IIC, 2604998 RPL3 ribosomal protein L3 Benign 'Suspicious HA & HC, 296423 RRAGD Ras-related GTP binding D Bcnign/Suspicious succinate dehydrogenase complex, subunit A, HA & HC, 2798.538 SDHA flavoprotein (Fp) Benign.Suspicious Succinate dehydrogenase complex, Subunit A, IIA & IIC, 2798.538 SDHALP flavoprotein pseudogene 1 Benign.Suspicious succinate dehydrogenase complex, subunit A, HA & HC, 2798.538 SDHALP2 flavoprotein pseudogene 2 Benign.Suspicious Patent Application Publication Dec. 22, 2011 Sheet 9 of 28 US 2011/0312520 A1 FIGURE 4D TCD GENE Gene Description Classificatio Pahel succinate dehydrogenase complex, subunit A. HA & HC, 2798.538 SDAP3 flavoprotein pseudogene 3 Benign/Suspicious solute carrier family l6, member 1 (monocarboxylic FIA & HC, 24285O1 SLC16A1 acid transporter 1) Benign/Suspicious IIA & C. 28775.08 SNORD63 - small nucleolar RNA C/D box 63 Benign/Suspicious HA & HC, 25.62529 ST3GAL5 ST3 beta-galactoside alpha-2,3-sialyltransferase 5 Benign.Suspicious HA & HC, 2688499 ZBED2 zinc finger, BED-type containing 2 Benign.Suspicious ATP-binding cassettic, sub-family D (ALD), member 3450861 ABCD2 2. ------------------------------------------------------------ Benign Suspicious 334106 ACER3 alkaline ceramidase 3 Benign.Suspicious 27.96553 ACSL acyl-CoA synthetase long-chain family member 1 Benign.Suspicious 3375735 AHNAK AHNAK nucleoprotein Benign"Suspicious 2439554 AIM2 absent in melanoma 2 Benign.Suspicious 37684.74 ARSG arylsulfatase G Benign.Suspicious 32.14845 ASPN asporin Benign.Suspicious 3OO6572 AUTS2 autism susceptibility candidate 2 Benign.Suspicious 3902489 BCL2L1 BCL2-like 1 Benign/Suspicious 268877 | BTLA B and T lymphocyte associated Benign.Suspicious 2708855 C11orf72