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  • 1. Recombinant XTEN Products XTEN fused to Human Proteins or Peptides • Long, tunable PK enables up to once-a-month dosing • Long PK maximizes efficacy and minimizes toxicity • Clinically validated in 2 products Protein or • Genetic fusion (no chemical conjugation step) DNA Peptide XTEN • Manufactured in E. coli or mammalian cells Expression • Non-immunogenic • Biodegradable (safer than PEG) Payload XTEN • 3 publications Products with Completed POC Trials • Exenatide-XTEN Diartis Inc. Ph.I in patients completed Long tail of natural hydrophilic amino acids • hGH-XTEN Versartis Inc. Ph.I in patients completed Preclinical Products • GLP2-XTEN Short Bowel Syndrome; Pre-clinical • AAT-XTEN AAT deficiency, Diabetes; Preclinical • Glucagon-XTEN 4 Diabetes indications; Preclinical • C-peptide-XTEN Diabetes complications; Preclinical1
  • 2. Advantages of Long Half-life Dosing Frequency Cmax: Toxicity Serum Concentration of Drug Toxic dose Therapeutic Window Peak-Trough Ratio: Toxicity Therapeutic dose AUC: Efficacy Time • Maximizes Efficacy (AUC) • Improves Dosing Frequency • MinimizesToxicity (Cmax, P/T ratio) • Improves Compliance2
  • 3. XTEN Concept O O O O O O O PEG O O O N N N N N N N Polypeptide O O O O O O O N XTEN N Drug N N N N N O O O O Promoter O XTEN PEG and XTEN are linear polymers with large apparent molecular weight3
  • 4. PEG versus XTEN DNA Drug DNA Drug XTEN Expression Expression GMP PEG XTEN K Unstructured polymer with large hydrodynamic radius K K K N K + Half-life extension + Half-life extension (Tunable) + Low immunogenicity + Low immunogenicity  Complicated manufacturing + Improved manufacturing  Heterogeneous product + Homogeneous product4
  • 5. XTEN Design and Properties Goals Long Serum Half-life Stable in Plasma Composition G, E, S, T, A, P Degraded Intracellularly Size 864 AA = 75 kDa High Expression Level Expression E. coli & Mammalian Genetically Stable No Non-specific Binding SDS-PAGE, SEC, MS Homogeneous No Aggregation Solubility >130 mg/ml Non-Immunogenic Viscosity Same as PEG Solution Plasma Stability >30 days No unstable AA Biodegradable Yes No hydrophobic AA Endotoxin/DNA/HCP All easily removed Multiple AA types Not Repetitive Evolutionary Process Current XTEN meets all goals and requirements5
  • 6. XTEN Technology Natural Peptide Structure Monodisperse by ESI-MS …GSTSESPSGTAPGTSPSGESSTAP GSTSESPSGTAPGSTSESPSGTAPGT STPESGSASPGTSTPESGSASPGST SESPSGTAPGSTSESPSGTAPGTSPS GESSTAPGSTSESPSGTAPGTSPSG ESSTAPGTSPSGESSTAPGSTSSTAE SPGPGTSPSGESSTAPGTSPSGESS TAPGSTSE… Unstructured by CD Large Hydrodynamic Radius by SEC XTEN XTEN is a polymer with similar properties as PEG, but is a protein6 Schellenberger et al., (2009) Nature Biotechnology, 27, 1186-90
  • 7. XTEN Immunogenicity Summary - Zero hydrophobic residues - Zero epitopes by Tepitope prediction - Zero epitopes by Epivax prediction - Morphosys panned 4 times – no binders - Non-immunogenic in in-house animal studies - Mice, rats, rabbit, cyno: 439 epitopes; 67 animals; <10 sc injections - Non-immunogenic in 2 toxicology studies - Mice: 14 SC injections; Monkeys: 5 SC injections - Monkeys: 7 SC injections - Non-immunogenic in 2 clinical trials (120 patients) - 1 monthly dose, but Exenatide payload shows response after 10 days - Non-immunogenic even with Freund’s adjuvant - 20 mice, 6 rabbits: antibody titer too low to be useful • All data suggest that XTEN is non-immunogenic in animals • Because XTEN is as foreign to animals as to humans, XTEN is expected to be non-immunogenic in humans7 • Initial clinical results support this conclusion
  • 8. Immunogenicity of XTEN Response XTEN N Species Injections XTEN Control Length XTEN_A 10 mouse 10, sc weak strong 576 - 10 different XTENs XTEN_A 3 mouse 9, sc none strong 576 - All 100% G,E,S,T,A,P XTEN_A 3 cyno 3, iv none NA 576 XTEN_C 4 rabbit 3, sc none weak 288 - Different ratios XTEN_C 4 mouse 3, sc none weak 288 - Comprising 436 9-mers XTEN_C 4 rat 3, sc none weak 288 - Non-repetitive XTEN_F 3 mouse 9, sc none strong 875 - No T-cell epitopes XTEN_G 3 rat 4, sc/ip none strong 288 - Immunized 67 animals XTEN_H 3 rat 4, sc/ip none strong 288 - 4 species XTEN_I 3 rat 4, sc/ip none strong 288 - Up to 10 SC injections XTEN_K 3 mouse 9, sc none strong 875 XTEN_L 5 mouse 9, sc none strong 144 XTEN_L 3 mouse 3, sc none strong 144 Immune Response Rating: Weak < 0.3OD at 1:100 XTEN_M 5 mouse 9, sc weak strong 144 Moderate >1.0OD at 1:100; XTEN_M 3 mouse 3, sc none strong 144 <0.3OD at 1:1000 XTEN_N 5 mouse 9, sc none strong 144 Strong > 0.3OD @ 1:1000 XTEN_N 3 mouse 3, sc none strong 144 • No significant immune responses to any XTEN sequences tested • No response to the clinical XTEN sequence • Even with adjuvant generally no immune response8
  • 9. XTEN Safety Advantages over PEG • PEG is not biodegraded • PEGylated proteins can accumulate in the kidneys, causing vacuolation Bendele, A., et al. (1998) Toxicol Sci, 42: 15 • Products are routinely tested for vacuolation – and have failed for this reason • “The non-biodegradable nature of PEG may become a limiting factor for the next generation of protein pharmaceuticals” Gaberc-Porekar, V., et al.(2008) Curr Opin Drug Discov Devel, 11: 242 • XTEN is fully biodegradeable • Stable in serum, but rapidly degraded by intracellular and lysosomal proteases • No kidney vacuolation (or other toxicities) observed • Single dose of 200 mg/kg • Multiple doses of 50 mg/kg The accumulation of PEG in all tissues has become a serious concern9
  • 10. Biodegradation of XTEN vs PEG • PEG is not biodegraded • PEGylated proteins can accumulate in the kidneys, causing vacuolation Bendele, A., et al. (1998) Toxicol Sci, 42: 15 • Products are routinely tested for vacuolation – and have failed for this reason • XTEN is fully biodegradeable • Stable in serum, but rapidly degraded by lysosomal proteases • No kidney vacuolation (or other toxicities) observed • Single dose of 200 mg/kg; Multiple doses of 50 mg/kg In Monkey Plasma In Monkey In vivo In Kidney Homogenate Time (days) Time (days) Time (days) 0 1 7 0 1 7 0 1 7 Detection: Anti-GFP Western of blood samples Incubated @ 37C in 50% plasma. Injected into a cynomolgus monkey Incubated @ 37C in rat kidney homogenate XTEN is stable in blood, but rapidly degraded intracellularly, as desired10
  • 11. Process Advantage over PEG XTEN PEG GLP2-XTEN880 G-CSF-PEG20 (Neulasta) Calc. 84,830 Da Exp. 84,831 DaIntensity82000 84000 86000 88000 Mass Mass Bagal, D., et al. (2008) Anal. Chem., 80: 2408-18 1 Species >100 Species Homogeneity of XTEN is a major advantage11
  • 12. 6-Month Stability of XTEN -80’C, 4’C and 25’C SEC 670kD 44kD 17kD 150kD 1.3kD SDS-PAGE Temp (°C) -80 4 25 37 37’C -80’C, 4’C RPC18 25’C 37’C Degraded XTEN 37’C 25’C XTEN is stable in water for 6 months at 25C, pH 6.212
  • 13. Lot Release MethodsProperty Methods Performed at Amunix StatusPurity SDS-PAGE Developed Reverse Phase HPLC – Intact protein Developed Analytical SEC Developed Analytical IEX DevelopedConcentration A280 DevelopedIdentity Intact Mass Determination – ESI mass spectrometry Developed Peptide Mapping – Tryptic digest, RP-C18 and MALDI DevelopedSafety Endotoxin – Endosafe cartridges Developed Host Cell Protein – Cygnus ELISA kit Developed Host Cell DNA – qPCR DevelopedPotency Payload Specific assays: enzymatic, cell based, binding Developed Outsourced Methods: CD, MALS, AA sequence, AA composition13
  • 14. GLP2-XTEN PK in Cynos: SC vs IV 2 mg/kg, 3 cynos/group; XTEN/GLP2 ELISA IV = 118 hrs SC = 125 hrs ng/ml hours Cyno T½ is ~125 hr Bioavailability is 93%14
  • 15. Cyno PK of AAT-XTEN Alpha1-anti-Trypsin-XTEN, IV, 2 cynos/groupConcentration (ng/ml) T1/2= 130 hrs T1/2= 111 hrs Time (hours) Cyno T1/2 at expected dosing is 130 hrs15
  • 16. hGH-XTEN PK in Cynos: SC, IV, IM 1000000 2 cynos/arm 1.5 mg/kg SC 1.5 mg/kg IV 1.5 mg/kg 1.5 mg/kg IM Total Protein Conc. (ng/ml) 100000 10000 IM T1/2 = 110 hrs SC IV 1000 100 0 50 100 150 200 250 300 350 400 Time (hours) IV, SC and IM are nearly identical - very high bioavailability16
  • 17. Allometric Scaling of XTEN PK 300 XTEN Product GLP2-XTEN Ex4 hGH GLP2G (267 hrs) 250 aaT FVII FIX 200 Half-Life (hours) Exenatide-XTEN 150 130 hrs 100 50 0 Mouse Rat Monkey Dog Human XTEN extends half-life proportional to species size17
  • 18. XTEN Publications18